![4 Rancour, Backues, and Bednarek
expression. Taking the following details into consideration while
determining the best antigen to express will greatly increase the
chance that quality antibodies will be generated.
The first consideration is whether the protein of interest is
known or predicted to be a soluble, a peripheral membrane, or
an integral membrane protein. Soluble proteins or domains are
much easier to work with in terms of expression, purification,
and, in some case, immunization protocols. For peripheral and
integral membrane proteins, select cDNA fragments predicted
to encode hydrophilic soluble domains of the protein of inter-
est (i.e., domains lacking long contiguous hydrophobic amino
acid stretches or transmembrane segments as predicted by hydro-
pathicity analysis using the Kyte–Doolittle (1) and/or Hopp–
Woods (2) algorithms or utilizing newer methods of soluble pro-
tein prediction such as the method of Smialowski et al. (3)).
Insoluble proteins can also be expressed and enriched as inclusion
bodies in E. coli and then subsequently used for immunization.
However, soluble proteins or protein domains most likely best
represent the native folding organization of the protein in its cel-
lular context and thus would be the optimal antigen to produce
antibodies that would be used for immunocytochemistry. Addi-
tional drawbacks to using insoluble proteins include the purity of
the protein not being high enough to generate specific anti-sera
against the antigen, and affinity purification of specific antibod-
ies becomes problematic (but not impossible) using the original
antigen. Insoluble proteins can limit the use of standard liquid
chromatography purification methods sans attempts at denatu-
ration and renaturation procedures. Therefore, soluble proteins
or domains should be the first choice for easy antigen produc-
tion and purification to ensure a clean antigen to induce a specific
immunological response.
The second consideration is whether the protein of interest is
post-translationally modified. If so, does this modification influ-
ence its functionality and/or localization? Proteins from eukary-
otes are commonly post-translationally modified. For example, in
cases where these proteins are extensively decorated with carbohy-
drates, these modifications may adversely influence the reactivity
of antibodies raised against an unmodified protein expressed in
E. coli. Conversely, if you are interested in antibodies against anti-
gens with specific modifications, you will need to keep in mind
that most eukaryotic cell modifications do not take place in E.
coli and thus you will need to seek other methods for antigen
production.
The third consideration is whether the protein of interest is
either a homo- or hetero-oligomer. Oligomerization may influ-
ence the ability to express the protein of interest as a soluble pro-
tein. A self-assembling soluble homo-oligomer [i.e., AtCDC48A
(4)] is much easier to produce than a subunit of a multimeric,](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-20-320.jpg)
![6 Rancour, Backues, and Bednarek
expressed protein for use as an immunogen, the resulting clones
must first be tested via small-scale induction to verify expression,
and then subsequently by small-scale fractionation to determine
the solubility of the protein product. Once the expression and
solubility of the protein has been verified, large-scale expression
and purification of the soluble protein by affinity chromatography
using glutathione resin can be initiated. GST tag removal from
purified soluble fusion proteins using sequence-specific proteases
followed by protease removal by affinity chromatography is the
final step in preparing a purified soluble bacterially expressed pro-
tein for use as an immunogen. For insoluble proteins, inclusion
bodies are isolated and used for immunization. In this chapter,
we present protocols for the preparation of tag-free soluble and
insoluble bacterially expressed proteins for use as immunogens.
2. Materials
2.1. SDS-PAGE
(Sodium
Dodecylsulfate-
Polyacrylamide Gel
Electrophoresis)
1. Precast Ready Gel Tris–HCl SDS-PAGE gels, 4–15% acry-
lamide, Tris/glycine/SDS buffer (Bio-Rad Laboratories,
Inc., Hercules, CA).
2. Small vertical electrophoresis unit (Hoefer, Inc., San Fran-
cisco, CA).
3. Power Supply (Bio-Rad Laboratories, Inc., Hercules, CA).
4. 10× SDS-PAGE running buffer: Mix 30.3 g Tris base,
144 g glycine, 10 g SDS, and distilled water (dH2O) to
a final volume of 1 L; dilute with dH2O to 1× strength
for use.
5. 5× SDS-PAGE sample buffer (5× SSB): Mix 3.9 mL 2 M
Tris–HCl pH 6.8, 2.5 g SDS, 12.5 mL glycerol, 8 mg bro-
mophenol blue, 6.25 mL β-mercaptoethanol, and double
distilled water (ddH2O) to a final volume of 25 mL. Use at
2× strength (2× SSB): 4 mL 5× SSB diluted with ddH2O
to final volume of 10 mL.
6. Protein Molecular Weight Markers: SDS-PAGE broad
range standards (Bio-Rad Laboratories, Inc., Hercules,
CA) diluted 1:40 in 2× SSB.
7. Coomassie stain for protein gels: 0.1% (w/v) Coomassie R-
250 dissolved in fixative [40% (v/v) methanol, 10% (v/v)
glacial acetic acid]; store capped at room temperature.
8. Coomassie destain solution for protein gels: 40% (v/v)
methanol, 10% (v/v) glacial acetic acid; store capped at
room temperature.
9. Kimwipes (Kimberly-Clark Corporation, Irving, TX).
10. Gel drying kit (Promega Corp., Madison, WI).](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-22-320.jpg)
![unless I be naked, Mistress whore!"[165]
So she loosed her petticoat-
trousers and, putting her hand under her clothes, tore them off her
body; then twisted up a silken kerchief into cord-shape, girt it round
her middle and became as she were a scald-head Ifritah or a spotted
snake. With this she inclined towards the damsel and said, "Do thou
as I have done." All this time, Sharrkan was gazing at the twain, and
laughing at the beldam's loathly semblance. So the damsel leisurely
rose and, taking a sash of Yamani stuff, passed it twice round her
waist, then she tucked up her trousers and displayed two calves of
alabaster carrying a mound of crystal, smooth and rounded, and a
stomach which exhaled musk from its dimples, as it were a bed of
Nu'uman's anemones; and breasts like double pomegranates. Then
the old woman leant towards her, and the two laid hold either of
each, while Sharrkan raised his head Heavenwards and prayed Allah
that the belle might beat the beldam. Presently the young woman
gat beneath the old woman; and, gripping her waist-cloth with the
left and circling her neck with the right hand, hoisted her off the
ground with both; whereupon the old woman strove to free herself
and, in so doing fell on her back arsiversy, with her legs high in air
and her hairy bush between them showed manifest in the
moonshine; furthermore she let fly two great farts[166]
one of which
blew up the dust from the earth's face and the other steamed up to
the gate of Heaven. Sharrkan laughed till he fell back upon the
ground. Then he arose and, baring his brand looked right and left,
but he saw no one save the old woman sprawling on her back, and
said to himself, "He lied not who named thee Lady of Calamities!
Verily thou knewest her prowess by her performance upon the
others." So he drew near them to hear what should pass between
them. Then the young lady went up to the old one and, throwing a
wrapper of thin silk upon her nakedness, helped her to don her
clothes and made excuses saying, "O my lady Zat al-Dawahi, I
intended only to throw thee and not all this, but thou triedst to twist
out of my hands; so laud to Allah for safety!" She returned her no
answer, but rose in her shame and walked away till out of sight,
leaving the handmaids prostrate and pinioned, with the fair damsel
standing amongst them. Quoth Sharrkan to himself, "Every luck hath](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-27-320.jpg)
![its cause. Sleep did not fall upon me nor the war-horse bear me
hither save for my good fortune; for doubtless this maid and what is
with her shall become booty to me." So he made towards his steed
and mounted and heeled[167]
him on, when he sped as the shaft
speeds from the bow, and in his hand he still hent his brand bare of
sheath, which he brandished shouting the while his war-cry, "Allah is
All-mighty[168]
!" When the damsel saw him she sprang to her feet
and, taking firm stand on the bank of the stream, whose breadth
was six ells, the normal cubits, made one bound and landed clear on
the farther side,[169]
where she turned and cried out with a loud
voice, "Who art thou, O thou fellow, that breakest in upon our
privacy and pastime, and that too hanger in hand as if charging a
host? Whence camest thou and whither art thou going? Speak
sooth, for truth will stand thee in good stead, and lie not, for lies
come of villein-breed. Doubtless thou hast wandered this night from
thy way, that thou chancedst upon this place whence escape were
the greatest of mercies; for thou art now in an open plain and, did
we shout but a single shout, would come to our rescue four
thousand knights.[170]
So tell me what thou wantest; and if thou
wouldst only have us set thee on the right road, we will do so."
When Sharrkan heard her words he replied, "I am a stranger of the
Moslems, who fared forth this night single-handed, seeking for spoil;
nor could this moonlight show me a fairer booty than these ten
maidens; so I shall seize them and rejoin my comrades with them."
Quoth she, "I would have thee know that as for the booty thou hast
not come at it; and, as for the handmaids, by Allah, they shall never
be thy spoil. Have I not told thee that to lie is villein-vile?" Quoth he,
"The wise man is he who taketh warning by others." Thereupon
quoth she, "By the truth of the Messiah, did I not fear that thy death
would be on my hands, I would shout a shout should fill the mead
for thee with war steeds and with men of might, but I take pity upon
the stranger. So, if thou seek booty, I require of thee that thou alight
from thy steed and swear to me, by thy faith, that thou wilt not
advance against me aught like arms in hand, and we will wrestle, I
and thou. If thou throw me, set me on thy steed and take all of us
to thy booty; but if I throw thee, thou shalt become under my](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-28-320.jpg)
![command. Swear this to me, for I fear thy treachery: indeed it hath
become a common saw, Where Perfidy is innate there Trust is a
weakly mate. Now an thou wilt swear I will return and draw near to
thee and tackle thee." Answered Sharrkan (and indeed he lusted to
seize her and said in his soul, "Truly she knoweth not that I am a
champion of champions"); "Swear me by what oath thou wilt and by
what thou deemest most binding, and I will not approach thee with
aught till thou hast made thy preparation and sayest:—Draw near
that I wrestle with thee. If thou throw me, I have money
wherewithal to ransom myself; and if I throw thee, 'twill be booty
and booty enough for me!" Rejoined the damsel, "I am content
herewith!" and Sharrkan was astounded at her words and said, "And
by the truth of the Apostle (whom Allah bless and keep!) I too am
content on the other part!" Then said she, "Swear to me by Him who
sprite in body dight and dealt laws to rule mankind aright, that thou
wilt not offer me aught of violence save by way of wrestling; else
mayst thou die without the pale of Al-Islam." Sharrkan replied, "By
Allah! were a Kazi to swear me, even though he were a Kazi of the
Kazis,[171]
he would not impose upon me such an oath as this!" Then
he sware to her by all she named and tied his steed to a tree; but he
was drowned in the sea of thought, saying in himself, "Praise be to
Him who fashioned her from dirty water!"[172]
Then he girt himself
and made ready for wrestling, and said to her, "Cross the stream to
me;" but she replied, "It is not for me to come over to thee: if thou
wilt, pass thou over here to me." "I cannot do that," quoth he, and
quoth she, "O boy, I will come across to thee." So she tucked up her
skirts and, leaping, landed on the other side of the stream by his
side; whereupon he drew near to her and bent him forwards and
clapped palms.[173]
But he was confounded by her beauty and
loveliness; for he saw a shape which the Hand of Power had tanned
with the dye-leaves of the Jánn, which had been fostered by the
Hand of Beneficence and fanned by the Zephyrs of fair fortune and
whose birth a propitious ascendant had greeted. Then she called out
to him, "O Moslem, come on and let us wrestle ere the break of
morning," and tucked up her sleeves from a fore-arm like fresh curd,
which illumined the whole place with its whiteness; and Sharrkan](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-29-320.jpg)
![the ground. He fell on his back,[174]
and then she said to him, "Rise:
I give thee thy life a second time. I spared thee in the first count
because of thy Prophet, for that he made unlawful the slaying of
women; and I do so on the second count because of thy weakliness
and the greenness of thine years and thy strangerhood; but I charge
thee, if there be in the Moslem army sent by Omar bin al-Nu'uman
to succour the King of Constantinople, a stronger than thou, send
him hither and tell him of me: for in wrestling there are shifts and
trips, catches and holds, such as the feint or falsing and the snap or
first grip, the hug, the feet-catch, the thigh-bite,[175]
the jostle and
the leg-lock." "By Allah, O my lady," quoth Sharrkan (and indeed he
was highly incensed against her), "had I been Master al-Safdí,
Master Mohammed Kimál or Ibn al-Saddí,[176]
as they were in their
prime, I had kept no note of these shifts thou mentionest; for O my
mistress, by Allah, thou hast not grassed me by thy strength, but by
the blandishments of thy back-parts; for we men of Mesopotamia so
love a full-formed thigh that nor sense was left me nor foresight. But
now, an thou wish, thou shalt try a third fall with me while my wits
are about me, and this last match is allowed me by the laws of the
game which sayeth the best of three: moreover I have regained my
presence of mind." When she heard his words she said to him, "Hast
thou not had a belly-full of this wrestling, O vanquished one?
However come on, an thou wilt; but know that this must be the last
round." Then she bent forward and challenged him and Sharrkan did
likewise, setting to it in real earnest and being right cautious about
the throw: so the two strove awhile and the damsel found in him a
strength such as she had not observed before and said to him, "O
Moslem, thou art now on thy mettle." "Yes," he replied, "thou
knowest that there remaineth to me but this one round, after which
each of us will wend a different way." She laughed and he laughed
too;[177]
then she overreached at his thigh and caught firm hold of it
unawares, which made him greet the ground and fall full on his
back. She laughed at him and said, "Art thou an eater of bran? Thou
art like a Badawi's bonnet which falleth off with every touch or else
the Father of Winds[178]
that droppeth before a puff of air. Fie upon
thee, O thou poor thing!" adding, "Get thee back to the Moslem](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-31-320.jpg)
![army and send us other than thyself, for thou failest of thews; and
proclaim for us, among the Arabs and Persians, the Turks and
Daylamites,[179]
whoso hath might in him, let him come to us." Then
she made a spring and landed on the other side of the stream and
said to Sharrkan, laughing, "Parting with thee is right grievous to
me, O my lord; but get thee to thy mates before dawn, lest the
Knights come upon thee and pick thee up on their lance-points.
Thou hast no strength to defend thee against a woman, so how
couldst thou hold thine own amongst men of might and Knights?"
Sharrkan was confounded and called to her (as she turned from him
making towards the convent), "O my lady, wilt thou go away and
leave the miserable stranger, the broken hearted slave of love?" So
she turned to him laughing and said, "What is thy want? I will grant
thee thy prayer." "Have I set foot in thy country and tasted the
sweetness of thy courtesy," replied he, "and shall I return without
eating of thy victual and tasting thy hospitality; I who have become
one of thy servitors!" "None baulk kindliness save the base," she
rejoined, "honour us in Allah's name, on my head and eyes be it!
Mount thy steed and ride along the brink of the stream over against
me, for now thou art my guest." At this Sharrkan was glad and,
hastening back to his horse, mounted and walked him abreast of
her, and she kept faring on till they came to a drawbridge[180]
built of
beams of the white poplar, hung by pullies and steel-chains and
made fast with hooks and padlocks. When Sharrkan looked, he saw
awaiting her upon the bridge the same ten handmaids whom she
had thrown in the wrestling-bouts; and, as she came up to them,
she said to one in the Greek tongue, "Arise and take the reins of his
horse and conduct him across into the convent." So she went up to
Sharrkan and led him over, much puzzled and perturbed with what
he saw, and saying to himself, "O would that the Wazir Dandan were
here with me that his eyes might look upon these fairest of favours."
Then he turned to the young lady and said to her, "O marvel of
loveliness, now I have two claims upon thee; first the claim of good-
fellowship, and secondly for that thou hast carried me to thy home
and offered me thy hospitality. I am now under thy commandance
and thy guidance; so do me one last favour by accompanying me to](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-32-320.jpg)
![the lands of Al-Islam; where thou shalt look upon many a lion-
hearted warrior and thou shalt learn who I am." When she heard
this she was angered and said to him, "By the truth of the Messiah,
thou hast proved thyself with me a man of keen wit; but now I see
what mischief there is in thy heart, and how thou canst permit
thyself a speech which proveth thy traitorous intent. How should I
do as thou sayest, when I wot that if I came to that King of yours,
Omar bin al-Nu'uman, I should never get free from him? For truly he
hath not the like of me or behind his city walls or within his palace-
halls, Lord of Baghdad and of Khorasan though he be, who hath
built for himself twelve pavilions, in number as the months of the
year, and in each a concubine after the number of the days; and if I
come to him he would not prove shy of me, for your folk believe I
am lawful to have and to hold as is said in your writ:—Or those
women whom your right hand shall possess as slaves.[181]
So how
canst thou speak thus to me? As for thy saying:—Thou shalt look
upon the braves of the Moslems, by the truth of the Messiah, thou
sayest that which is not true, for I saw your army when it reached
our land, these two days ago; and I did not see that your ordinance
was the ordinance of Kings, but I beheld only a rabble of tribesmen
gathered together. And as to thy words:—Thou shalt know who I
am, I did not do thee kindness because of thy dignity but out of
pride in myself; and the like of thee should not talk thus to the like
of me, even wert thou Sharrkan, Omar bin al-Nu'uman's son, the
prowest name in these days!" "Knowest thou Sharrkan?" asked he;
and she answered "Yes! and I know of his coming with an army
numbering ten thousand horsemen; also that he was sent by his sire
with this force to gain prevalence for the King of Constantinople." "O
my lady," said Sharrkan, "I adjure thee by thy religion, tell me the
cause of all this, that sooth may appear to me clear of untruth, and
with whom the fault lies." "Now by the virtue of thy faith," she
replied, "did I not fear lest the news of me be bruited abroad that I
am of the daughters of Roum, I would adventure myself and sally
forth single-handed against the ten thousand horsemen and slay
their leader, the Wazir Dandan and vanquish their champion
Sharrkan.[182]
Nor would aught of shame accrue to me thereby, for I](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-33-320.jpg)
![enow), "Verily if Sharrkan fell into my hands, I would go forth to him
in the habit of a man and drag him from his saddle-seat and make
him my captive and lay him in bilboes," pride and passion and
knightly jealousy took possession of him and he desired to discover
and declare himself and to lay on load; but her loveliness restrained
him and he began repeating:—
An faulty of one fault the Beauty prove, ✿ Her charms a thousand advocates shall
move.
So she went up and Sharrkan after her; and, when he saw the
maiden's back and hinder cheeks that clashed against each other,
like rollers in the rolling sea, he extemporised these couplets:—
For her sins is a pleader that brow, ✿ And all hearts its fair pleading must trow:
When I saw it I cried, "To-night ✿ The moon at its fullest doth show;
Tho' Balkís' own Ifrit[183] try a bout, ✿ Spite his force she would deal him a throw.
The two fared on till they reached a gate over which rose a marble
archway. This she opened and ushered Sharrkan into a long
vestibule, vaulted with ten connected arches, from each of which
hung a crystal lamp glistening like a spark of fire. The handmaids
met her at the further end bearing wax candles of goodly perfume,
and wearing on their heads golden fillets crusted with all manner
bezel-gems,[184]
and went on before her (Sharrkan still following), till
they reached the inner convent. There the Moslem saw couches and
sofas ranged all around, one opposite the other and all overhung
with curtains flowered in gold. The monastery floor was paved with
every kind of vari-coloured marbles and mosaic-work, and in the
midst stood a basin that held four-and-twenty jetting fountains of
gold, whence the water ran like molten silver; whilst at the upper
end stood a throne spread with silks fit only for Kings. Then said the
damsel, "Ascend, O my lord, this throne." So he went up to it and
sat down and she withdrew to remain absent for some time.
Sharrkan asked of her from one of the servants who answered him,
"She hath gone to her dormitory; but we will serve thee even as she
ordered." So they set before him viands of rare varieties, and he ate](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-36-320.jpg)
![his sufficiency, when they brought him a basin of gold and an ewer
of silver, and he washed his hands. Then his thoughts reverted to his
army, knowing not what had befallen it in his absence and calling to
mind also how he had forgotton his father's injunctions: so he was
troubled about his case, repenting of what he had done till the dawn
broke and the day appeared; when he lamented and sighed and
became drowned in the sea of sadness and repeated:—
I am not lost to prudence, but indeed ✿ Here I'm bewildered, what shall be my
rede?
Would any aid me in mine ails of love, ✿ By my own might and sleight would I be
free'd:
But ah! my heart is lost and passion-shent: ✿ To none save Allah can I trust my
need!
When he ended his verse behold, there came up to him a rare show
and a fair, more than twenty maidens like crescents encompassing
the young lady, who shone in their midst as the full moon among the
constellations guarding and girding her. She was clad in brocades
befitting Kings; her breasts were like twin pomegranates, a woven
zone set with all kinds of jewels tightly clasped her waist which
expanded below into jutting hips; and her hinder cheeks stood out
as a mound of crystal[185]
supporting a silvern shaft. When Sharrkan
looked at her his wits went nigh to fly away from him with delight;
and he forgot army and Wazir as he gazed on her fair head decked
and dight with a net-work of pearls set off by divers sorts of gems.
Handmaids on her right and handmaids on her left bore her train, as
she paced with dainty graceful gait in all the pride of seemlihead. He
sprang to his feet seeing such beauty and loveliness, and cried
aloud, "Beware and beware of that zone rarely fair!" and broke out
into these couplets:—](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-37-320.jpg)
![With heavy back-parts, high breasts delicate, ✿ And lissome form that sways with
swimming gait,
She deftly hides love-longing in her breast; ✿ But I may never hide its ban and
bate:
While hosts of followers her steps precede,[186] ✿ Like pearls now necklaced and
now separate.
She gazed upon him for a long time and considered him till she was
assured of him, when she came up to him and said, "In very sooth
the place is honoured and illumined by thee, O Sharrkan! How sped
thy night, O hero, after we went away and left thee?"; adding,
"Verily lying is a vile thing and a shameful, especially in great Kings!
and thou art Crown-Prince Sharrkan, son and heir of King Omar bin
al-Nu'uman; so henceforth make no secret of thy rank and condition,
nor let me hear aught from thee but the truth; for leasing
bequeatheth hate and despite. And as thou art pierced by the shaft
of Fate, be resignation thine and abide content to wait." When he
heard her words he saw that artifice availed him naught and he
acknowledged the truth, saying, "I am Sharrkan, bin Omar bin al-
Nu'uman, whom fortune hath afflicted, and cast into this place; so
whatso thou willest, do it in my case!" She hung her head
groundwards a long while, then turned to him and said, "Be of good
cheer and let thine eyes be cool and clear;[187]
for thou art the guest
of my hospitality, and bread-and-salt hath made a tie between me
and thee; wherefore thou art in my ward and under my safeguard.
Have no fear for, by the truth of the Messiah, if all on earth sought
to do thee hurt they should not come at thee, till life had left my
body for thy sake: indeed thou art now under the charge of the
Messiah and of me." Hereat she sat her down by his side and fell to
playing with him, till his alarm subsided and he knew that had she
desired to slay him, she would have done so during the past night.
Presently she bespoke in the Grecian tongue one of her slave-girls,
who went away and soon came back bringing a beaker and a tray of
food; but Sharrkan abstained from eating and said to himself, "Haply
she hath put somewhat in this meat." She knew what was in his
thought; so she turned to him and said, "By the truth of the](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-38-320.jpg)
![Messiah, the case is not on such wise, nor is there aught in this
meat of what thou suspectest! Had my mind been set on slaying
thee, I had slain thee ere now." Then she walked up to the tray and
ate of every dish a mouthful; whereupon Sharrkan came forward
and ate too. She was pleased at this and both ate till they were
satisfied. They washed their hands and after that she rose and
ordered a handmaid to bring perfumes and herbs of sweet savour,
wines of all colours and kinds and a wine-service with vessels of
gold, silver and crystal. She filled a first goblet and drank it off
before offering it to him, even as she had done with the food: then
she downed a second and handed it to him. He drank and she said
to him, "O Moslem, see how thou art here in all solace and delight of
life!" And she ceased not to drink and ply him with drink, till he took
leave of his wits,——And Shahrazad perceived the dawn of day, and
ceased saying her permitted say.
Now when it was the Forty-ninth Night,
She said, It hath reached me, O auspicious King, that the damsel
ceased not to drink and ply Sharrkan with drink till he took leave of
his wits, for the wine and the intoxication of love he bore her.
Presently she said to the slave-girl, "O Marjánah[188]
! bring us some
instruments of music!" "To hear is to obey," said the handmaid and
going out, returned in the twinkling of an eye with a Damascus lute,
[189]
a Persian harp, a Tartar pipe, and an Egyptian dulcimer. The
young lady took the lute and, after tuning each several string, began
in gentle undersong to sing, softer, than zephyr's wing and sweeter
than Tasmin[190]
-spring, with heart safe and secure from everything
the couplets following:—](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-39-320.jpg)
![Allah assain those eyne! What streams of blood they shed! ✿ How many an
arrowy glance those lids of thine have sped.
I love all lovers who to lovers show them dure; ✿ Twere wrong to rue the love in
wrong-head born and bred:
Haply fall hapless eye for thee no sleeping kens! ✿ Heaven help the hapless heart
by force of thee misled!
Thou doomest me to death who art my king, and I ✿ Ransom with life the
deemster who would doom me dead.
Thereupon each and every of the maidens rose up and taking an
instrument, played and recited couplets in the Roumi tongue; then
their mistress sang also and seeing Sharrkan in ecstasies asked him,
"O Moslem, dost thou understand what I say?"; and he answered,
"Nay, my ecstasy cometh from the beauty of thy finger-tips." She
laughed and continued, "If I sing to thee in Arabic what wouldst
thou do?" "I should no longer," quoth he, "be master of my senses."
Then she took an instrument and, changing the measure, began
singing these verses:—
The smack of parting's myrrh to me, ✿ How, then, bear patience' aloë?
I'm girt by ills in trinity ✿ Severance, distance, cruelty!
My freedom stole that fairest she, ✿ And parting irks me bitterly.
When she ended her verse, she looked at Sharrkan and found him
lost to existence, and he lay for a while stretched at full length and
prone among the maidens.[191]
Then he revived and, remembering
the songs, again inclined to mirth and merriment; and the twain
returned to their wine and wassail, and continued their playing and
toying, their pastime and pleasure till day ceased illuminating and
night drooped her wing. Then the damsel went off to her dormitory
and when Sharrkan asked after her they answered, "She is gone to
her sleeping-chamber," whereto he rejoined, "Under Allah's ward and
His good guard!" As soon as it was morning, a handmaid came to
him and said to him, "My mistress biddeth thee to her." So he rose
and followed her and, as he drew near her lodging, the damsels
welcomed him with smitten tabrets and songs of greeting, and led
him through a great door of ivory studded with pearls and jewels.](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-40-320.jpg)
![Thence they passed with him into a tall and spacious hall, at the
upper end of which was a wide daïs carpeted with all kinds of silks,
and round it open lattices commanding a view of trees and streams.
About the saloon were figures carved in human form, and fashioned
on such wise that the air passed through them and set in motion
musical instruments within, so that the beholder would fancy they
spoke.[192]
Here sat the young lady, looking at the figures; but when
she saw Sharrkan, she sprang to her feet and, taking him by the
hand, made him sit down by her side, and asked him how he had
passed the night. He blessed her and the two sat talking awhile till
she asked him, "Knowest thou aught touching lovers and slaves of
love?"; and he answered "Yes! I wot somewhat in verse on that
matter." "Let me hear it," quoth she, so he began quoting:—
Pleasure and health, good cheer, good appetite ✿ To Azzah, freest with our name
and fame!
By Allah! would I near her off she flies ✿ At tangent, granting less the more I
claim:
I dote on Azzah, but when clear I off ✿ My rivals, clears me too that dearest
dame;
Like wandering wight that chose for shade a cloud ✿ Which, ere siesta done, thin
air became.
When she heard this she said, "Verily Al-Kuthayyir[193]
was
conspicuous for sweet speech and chaste, and he was superlative in
his praise of Azzah when he sang (and she began to recite):—
"Did Azzah deal behest to Sun o' noon, ✿ The judge had 'judged her beauty's
bestest boon;
And girls who come to me and carp at her, ✿ God make their rosy cheeks her
sandal-shoon!"
"And indeed," quoth she, "'twas said that Azzah boasted exceeding
beauty and loveliness." Then she asked Sharrkan saying, "O Prince,
dost thou know aught of Jamíl's[194]
verses to Buthaynah? if so
repeat to us somewhat of them;" and he answered, "Yes, I know
them better than any;" whereupon he began repeating these
couplets:—](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-41-320.jpg)
!["Jamíl, in Holy war go fight!" to me they say: ✿ What war save fight for fair ones
would I e'er essay?
To me their every word and work are mere delight, ✿ And martyrs clepe I all they
slay in fight and fray:
An ask I, "O Buthaynah! what's this love, I pray, ✿ Which eats my heart? quoth
she "'Twill stay for ever and aye!"
And when I cry, "Of wits return some small display ✿ For daily use," quoth she,
"Far, far 'tis fled away!"
Thou seekst my death; naught else thy will can satisfy ✿ While I no goal espy
save thee and thee alway.
"Thou hast spoken right well," said she, "O King's son, and Jamíl
also spoke excellently well. But what would Buthaynah have done
with him that he saith in his hemistich:—
Thou seekst my death; naught else thy will can satisfy?"
"O my lady," quoth Sharrkan, "she willed to do him what thou willest
to do with me, and even that will not satisfy thee." She laughed at
his opportune reply and they ceased not carousing till Day put out
her light and Night came in darkness dight. Then she rose and went
to her dormitory and slept, while Sharrkan slept in his place till
morning dawned. As soon as he awoke, the handmaids came to him
with tabrets and other instruments of mirth and merriment, as wont;
and, kissing the ground between his hands, said to him, "Bismillah!
—in Allah's name—be so kind as to come[195]
: our mistress biddeth
thee to her presence!" So he rose and accompanied the slave-girls
who surrounded him, playing on tabrets and other instruments of
music, till they passed from that saloon into another and a yet more
spacious hall, decorated with pictured likenesses and figures of birds
and beasts, passing all description. Sharrkan marvelled at the art
and artifice of the place and began reciting:—](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-42-320.jpg)
![He pluckt fruits of her necklace in rivalry, ✿ And her breast-pearls that bedded in
gold-mine lie.
Pure water on silvern bars is her brow, ✿ And her cheeks show roses with rubies
vie:
Meseems in her eyne that the violet's hue ✿ Lies purpling set in the Ithmid's[196]
dye.
When the lady saw Sharrkan, she stood up to him in honour and,
taking his hand, seated him by her side and asked, "O son of King
Omar bin al-Nu'uman, hast thou any cunning in the game of chess?"
"Yes," he answered, "but do not thou with me as said the poet:—
I speak and longing love upties me and unties me; ✿ Till with her honey-dew of
inner lip she plies me:
I brought the chess-board and my liefest lover plays me ✿ With white and black,
[197] but black-cum-white ne'er satisfies me:
'Twas as if King for Castle I were fain to place me ✿ Till wilful loss of game atwixt
two queens surprise me:
And if I seek to read intent in eyes that eye me ✿ Oh man! that glance askance
with hint of wish defies me."
Then she brought the chess-board and played with him; but
Sharrkan, instead of looking at her moves, kept gazing at her fair
mouth, and putting knight in place of elephant and elephant[198]
in
stead of knight. She laughed and said to him, "If thy play be after
this fashion, thou knowest naught of the game." "This is only our
first," replied he, "judge not by this bout." When she beat him he
replaced the pieces in position and played again with her; but she
beat him a second time, a third, a fourth and a fifth. So she turned
to him and said, "Thou art beaten in everything;" and he replied, "O
my lady, how should one playing with the like of thee avoid being
beaten?" Then she bade bring food, and they ate and washed their
hands; after which the wine was set before them and they drank.
Presently, she took the dulcimer, for her hand was cunning in smiting
it, and she began repeating to an accompaniment these couplets:—](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-43-320.jpg)
![impregnable place! The news of him reached King Hardub, thy
father, by report of the ancient dame Zat al-Dawahi; and thy sire,
our sovereign, hath made sure that thou hast rendered good service
to the army of the Greeks by taking captive this ominous lion." When
she heard this, she looked at the Knight and asked him, "What be
thy name?" and he answered, "I am Masurah, son of thy slave
Mausúrah bin Káshardah, Knight of Knights." "And how?" quoth she,
"durst thou enter my presence without leave?" Quoth he, "O my
lady, when I came to the gate, none forbade me, neither
chamberlain nor porter, but all the doorkeepers rose and forewent us
as of wont; although, when others come, they leave them standing
at the gate while they ask permission to admit them. But this is not
a time for long talking, when the King is expecting our return with
this Prince, the scorpion-sting[199]
of the Islamitic host, that he may
kill him and drive back his men whither they came, without the bane
of battling with them." "These words be ill words," rejoined the
Princess, "and Dame Zat al-Dawahi lied, avouching an idle thing and
a vain, whereof she weeteth not the truth; for by the virtue of the
Messiah, this man who is with me is not Sharrkan, nor is he a
captive, but a stranger who came to us seeking our hospitality, and I
made him my guest. So even were we assured that this be Sharrkan
and were it proved to us that it is he beyond a doubt, I say it would
ill befit mine honour that I should deliver into your hands one who
hath entered under my protection. So make me not a traitor to my
guest and a disgrace among men; but return to the King, my father,
and kiss the ground before him, and inform him that the case is
contrariwise to the report of the Lady Zat al-Dawahi." "O Abrizah,"
replied Masurah, the Knight, "I cannot return to the King's majesty
without his debtor and enemy." Quoth she (and indeed she had
waxed very wroth), "Out on thee! Return to him with my answer,
and no blame shall befal thee!" Quoth Masurah, "I will not return
without him." Thereupon her colour changed and she exclaimed,
"Exceed not in talk and vain words; for verily this man had not come
in to us, were he not assured that he could of himself and single-
handed make head against an hundred riders; and if I said to him:—
Thou art Sharrkan, son of King Omar bin al-Nu'uman, he would](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-45-320.jpg)
![ten?" "That would be villeiny," said she; "Let one have at one."
When he heard this, he sprang to his feet and made for them with
his sword and battle-gear; and Masurah, the Knight, also sprang up
and bore down upon him. Sharrkan met him like a lion and delivered
a shoulder cut[200]
which clove him to the middle, and the blade
came out gleaming and glittering from his back and bowels. When
the lady beheld that swashing blow, Sharrkan's might was magnified
in her sight and she knew that when she overthrew him in the
wrestle it was not by her strength but by her beauty and loveliness.
So she turned to the Knights and said, "Take wreak for your chief!"
Thereupon out came the slain man's brother, a fierce and furious
Knight, and rushed upon Sharrkan, who delayed not, but smote him
also with the shoulder-cut and the sword came out glittering from
his vitals. Then cried the Princess, "O ye servants of the Messiah,
avenge your comrade!" So they ceased not charging down upon
him, one after one; and Sharrkan also ceased not playing upon them
with the blade, till he had slain fifty knights, the lady looking on the
while. And Allah cast a panic into the hearts of the survivors, so that
they held back and dared not meet him in the duello, but fell upon
him in a body; and he laid on load with heart firmer than a rock, and
smote them and trod them down like straw under the threshing-sled,
[201]
till he had driven sense and soul out of them. Then the Princess
called aloud to her damsels, saying, "Who is left in the convent?";
and they replied, "None but the gate-keepers;" whereupon she went
up to Sharrkan and took him to her bosom, he doing the same, and
they returned to the palace, after he had made an end of the melée.
Now there remained a few of the Knights hiding from him in the cells
of the monastery, and when the Princess saw this she rose from
Sharrkan's side and left him for a while, but presently came back
clad in closely-meshed coat of ring-mail and holding in her hand a
fine Indian scymitar. And she said, "Now by the truth of the Messiah,
I will not be a niggard of myself for my guest; nor will I abandon
him though for this I abide a reproach and a byword in the land of
the Greeks." Then she took reckoning of the dead and found that he
had slain fourscore of the Knights, and other twenty had taken to
flight.[202]
When she saw what work he had made with them she said](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-47-320.jpg)
![to him, "Allah bless thee, O Sharrkan! The Cavaliers may well glory
in the like of thee." Then he rose and wiping his blade clean of the
blood of the slain began reciting these couplets:—
How oft in the mellay I've cleft the array, ✿ And given their bravest to lions a
prey:
Ask of me and of them when I proved me prow ✿ O'er creation, on days of the
foray and fray:
When I left in the onslaught their lions to lie ✿ On the sands of the lowlands[203]
in fieriest day.
When he ended his verse, the Princess came up to him with smiles
and kissed his hand; then she doffed her hauberk and he said to her,
"O lady mine, wherefore didst thou don that coat of mail and bare
thy brand?" "To guard thee against these caitiffs,"[204]
she replied.
Then she summoned the gate-keepers and asked them, "How came
ye to admit the King's Knights into my dwelling without leave of
me?"; and they answered, "O Princess, it is not our custom to ask
leave of thee for the King's messengers, and especially for the chief
of his Knights." Quoth she, "I think ye were minded only to disgrace
me and murder my guest;" and bade Sharrkan smite their necks. He
did so and she cried to the rest of her servants, "Of a truth, they
deserved even more than that!" Then turning to Sharrkan, she said
to him, "Now that there hath become manifest to thee what was
concealed, thou shalt be made acquainted with my history. Know,
then, that I am the daughter of King Hardub of Roum; my name is
Abrizah and the ancient dame, yclept Zat al-Dawahi, is my
grandmother by the sword side. She it certainly is who told my
father of thee, and as surely she will compass a sleight to slay me,
more by token as thou hast slain my father's chivalry and it is noised
abroad that I have separated myself from the Nazarenes and have
become no better than I should be with the Moslems. Wherefore it
were wiser that I leave this dwelling while Zat al-Dawahi is on my
track; but I require of thee the like kindness and courtesy I have
shown thee, for enmity will presently befal between me and my
father on thine account. So do not thou neglect to do aught that I
shall say to thee, remembering all this betided me not save by](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-48-320.jpg)
![reason of thee." Hearing her words, Sharrkan joyed greatly; his
breast broadened and his wits flew from him for delight, and he
said, "By Allah, none shall come at thee, while life is in my bosom!
But hast thou patience to bear parting from thy parents and thy
people?" "Even so," she answered; and Sharrkan swore to her and
the two plighted their troth. Then said she, "Now is my heart at
ease; but there remaineth one other condition for thee." "What is
it?" asked he and she answered, "It is that thou return with thy host
to thine own country." Quoth he, "O lady mine, my father, King Omar
bin al-Nu'uman, sent me to wage war upon thy sire, on account of
the treasure he plundered from the King of Constantinople, and
amongst the rest three great jewels, noted givers of good fortune."
Quoth she, "Cheer thy heart and clear thine eyes: I will tell thee the
whole of the tale and the cause of our feud with the King of
Constantinople. Know that we have a yearly festival, hight the
Convent-Feast, whereat Kings from all quarters and the noblest
women are wont to congregate; thither also come merchants and
traders with their wives and families, and the visitors abide there
seven days. I was wont to be one of them; but, when there befel
enmity between us, my father forbade me to be present at the
festival for the space of seven years. One year, it chanced that
amongst the daughters of the great who resorted to the patron, as
was their custom, came a daughter of the King of Constantinople, a
beautiful girl called Sophia. They tarried at the monastery six days
and on the seventh the folk went their ways;[205]
but Sophia said, I
will not return to Constantinople save by water. So they equipped for
her a ship in which she embarked with her suite; and making sail
they put out to sea; but as they were voyaging behold, a contrary
wind caught them and drove the vessel from her course till, as Fate
and Fortune would have it, she fell in with a Nazarene craft from the
Camphor Island[206]
carrying a crew of five hundred armed Franks,
who had been cruising about a long time. When they sighted the
sails of the ship, wherein Sophia and her women were, they gave
chase in all haste and in less than an hour they came up with her,
when they laid the grappling-irons aboard her and captured her.
Then taking her in tow they made all sail for their own island and](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-49-320.jpg)
![were but a little distant from it when the wind veered round and,
splitting their sails, drove them on to a shoal which lies off our coast.
Thereupon we sallied forth and, looking on them as spoil driven to
us by Fate,[207]
boarded and took them; and, slaying the men, made
prize of the wreck, wherein we found the treasures and rarities in
question and forty maidens, amongst whom was the King's daughter,
Sophia. After the capture we carried the Princess and her women to
my father, not knowing her to be a daughter of King Afridun of
Constantinople; and he chose out for himself ten including her; and
divided the rest among his dependants. Presently he set apart five
damsels, amongst whom was the King's daughter, and sent them to
thy father, King Omar bin al-Nu'uman, together with other gifts, such
as broadcloth[208]
and woollen stuffs and Grecian silks. Thy father
accepted them and chose out from amongst the five girls Sophia,
daughter of King Afridun; nor did we hear more of her till the
beginning of this year, when her father wrote to my father in words
unfitting for me to repeat, rebuking him with menaces and saying to
him:—Two years ago, you plundered a ship of ours which had been
seized by a band of Frankish pirates in which was my daughter
Sophia, attended by her maidens numbering some threescore. Yet
ye informed me not thereof by messenger or otherwise; nor could I
make the matter public, lest reproach befal me amongst the Kings,
by reason of my daughter's honour. So I concealed my case till this
year, when I wrote to certain Frankish corsairs and sought news of
my daughter from the Kings of the Isles. They replied:—By Allah we
carried her not forth of thy realm; but we have heard that King
Hardub rescued her from certain pirates. And they told me the whole
tale. Then he added in the writing which he writ to my father:—
Except you wish to be at feud with me and design to disgrace me
and dishonour my daughter, you will, the instant my letter reacheth
you, send my daughter back to me. But if you slight my letter and
disobey my commandment, I will assuredly make you full return for
your foul dealing and the baseness of your practices.[209]
When my
father read this letter and understood the contents,[210]
it vexed him
and he regretted not having known that Sophia, King Afridun's
daughter, was among the captured damsels, that he might have sent](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-50-320.jpg)
![her back to her sire; and he was perplexed about the case because,
after so long a time, he could not send to King Omar bin al-Nu'uman
and demand her back from him, especially as he had lately heard
that Heaven had granted him boon of babe by this Sophia. So when
we pondered that truth, we knew that this letter was none other
than a grievous calamity; and my father found nothing for it but to
write an answer to King Afridun, making his excuses and swearing to
him by strong oaths that he knew not his daughter to be among the
bevy of damsels in the ship and setting forth how he had sent her to
King Omar bin al-Nu'uman, who had gotten the blessing of issue by
her. When my father's reply reached King Afridun he rose up and sat
down,[211]
and roared and foamed at the mouth crying:—What! shall
he take captive my daughter and even her with slave-girls and pass
her on from hand to hand sending her for a gift to Kings, and they
lie with her without marriage-contract? By the Messiah and the true
Faith, said he, I will not desist till I have taken my blood-vengeance
for this and have wiped out my shame; and indeed I will do a deed
which the chroniclers shall chronicle after me! So he bided his time
till he devised a device and laid notable toils and snares, when he
sent an embassy to thy father, King Omar, to tell him that which thou
hast heard: accordingly thy father equipped thee and an army with
thee and sent thee to King Afridun, whose object is to seize thee
and thine army to boot. As for the three jewels whereof he told thy
father when asking his aid, there was not one soothfast word in that
matter, for they were with Sophia, his daughter; and my father took
them from her, when he got possession of her and of her maidens,
and gave them to me in free gift, and they are now with me. So go
thou to thy host and turn them back ere they be led deep into, and
shut in by, the land of the Franks and the country of the Greeks; for
as soon as you have come far enough into their interior, they will
stop the roads upon you and there will be no escape for you till the
Day of retribution and retaliation. I know that thy troops are still
halting where thou leftest them, because thou didst order a three
days' rest; withal they have missed thee all this time and they wot
not what to do." When Sharrkan heard her words, he was absent
awhile in thought; then he kissed Princess Abrizah's hand and said,](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-51-320.jpg)
!["Praise be to Allah who hath bestowed thee on me and appointed
thee to be the cause of my salvation and the salvation of whoso is
with me! But 'tis grievous to me to part from thee and I know not
what will become of thee after my departure." "Go now to thine
army," she replied, "and turn them back, while ye are yet near your
own country. If the envoys be still with them, lay hands on them and
keep them, that the case may be made manifest to you; and, after
three days, I will be with you all and we will enter Baghdad
together." As he turned to depart she said, "Forget not the compact
which is between me and thee;" then she rose to bid[212]
him
farewell and embrace him and quench the fire of desire, so she took
leave of him and, throwing her arms round his neck, wept with
exceeding weeping, and repeated these verses:—
I bade adieu, my right hand wiped my tears away, ✿ The while my left hand held
her in a close embrace:
"Fearest thou naught," quoth she, "of shame?" I answered "Nay, ✿ The lover's
parting day is lover's worst disgrace."
Then Sharrkan left her and walked down from the convent. They
brought his steed, so he mounted and rode down-stream to the
drawbridge which he crossed and presently threaded the woodland
paths and passed into the open meadow. As soon as he was clear of
the trees he was aware of horsemen which made him stand on the
alert, and he bared his brand and rode cautiously; but as they drew
near and exchanged curious looks he recognized them and behold, it
was the Wazir Dandan and two of his Emirs. When they saw him and
knew him, they dismounted and saluting him, asked the reason of
his absence; whereupon he told them all that had passed between
him and Princess Abrizah from first to last. The Wazir returned
thanks to Almighty Allah for his safety and said,[213]
"Let us at once
leave these lands; for the envoys who came with us are gone to
inform the King of our approach, and haply he will hasten to fall on
us and take us prisoners." So Sharrkan cried to his men to saddle
and mount, which they did and, setting out at once, they stinted not
faring till they reached the sole of the valley wherein the host lay.](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-52-320.jpg)
![The Ambassadors meanwhile had reported Sharrkan's approach to
their King, who forthright equipped a host to lay hold of him and
those with him. But Sharrkan, escorted by the Wazir Dandan and the
two Emirs, had no sooner sighted the army, than he raised the cry
"March! March!" They took horse on the instant and fared through
the first day and second and third day, nor did they cease faring for
five days; at the end of which time they alighted in a well-wooded
valley, where they rested awhile. Then they again set out and stayed
not riding for five and twenty days which placed them on the
frontiers of their own country. Here, deeming themselves safe, they
halted to rest; and the country people came out to them with guest-
gifts for the men and provender and forage for the beasts. They
tarried there two days after which, as all would be making for their
homes, Sharrkan put the Wazir Dandan in command, bidding him
lead the host back to Baghdad. But he himself remained behind with
an hundred riders, till the rest of the army had made one day's
march: then he called "To horse!" and mounted with his hundred
men. They rode on two parasangs'[214]
space till they arrived at a
gorge between two mountains and lo! there arose before them a
dark cloud of sand and dust. So they checked their steeds awhile till
the dust opened and lifted, discovering beneath it an hundred
cavaliers, lion-faced and in mail-coats cased. As soon as they drew
within earshot of Sharrkan and his meiny they cried out to them,
saying, "By the virtue of John and Mary, we have won to our wish!
We have been following you by forced marches, night and day, till
we forewent you to this place. So dismount and lay down your arms
and yield yourselves, that we may grant you your lives." When
Sharrkan heard this, his eyes stood out from his head and his cheeks
flushed red and he said "How is it, O Nazarene dogs, ye dare enter
our country and overmarch our land? And doth not this suffice you,
but ye must adventure your selves and address us in such unseemly
speech? Do you think to escape out of our hands and return to your
country?" Then he shouted to his hundred horsemen, "Up and at
these hounds, for they even you in number!" So saying, he bared his
sabre and bore down on them, he and his, but the Franks met them
with hearts firmer than rocks, and wight clashed against wight, and](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-53-320.jpg)
![armed cap-à-pie and clad in a surcoat of gold stuff, riding on a grey-
white steed,[215]
and he had no hair on his cheeks. He urged his
charger on to the midst of the battle-plain and the two fell to
derring-do of cut and thrust, but it was not long before the Frank
foined the Moslem with the lance-point; and, toppling him from his
steed, took him prisoner and led him off crestfallen. His folk rejoiced
in their comrade and, forbidding him to go out again to the field,
sent forth another, to whom sallied out another Moslem, brother to
the captive, and offered him battle. The two fell to, either against
other, and fought for a little while, till the Frank bore down upon the
Moslem and, falsing him with a feint, tumbled him by a thrust of the
lance-heel from his destrier and took him prisoner. After this fashion
the Moslems ceased not dashing forwards, one after one, and the
Franks to unhorse them and take them captive, till day departed and
the night with darkness upstarted. Now they had captured of the
Moslems twenty cavaliers, and when Sharrkan saw this, it was
grievous to him and he mustered his men and said to them, "What is
this thing that hath befallen us? To-morrow, I myself will go forth to
the field and offer singular combat to their chief and learn what is
the cause of his entering our land and warn him against doing battle
with our band. If he persist, we will punish him with death, and if he
prove peaceable we will make peace with him." They nighted on this
wise till Allah Almighty caused the morn to dawn, when mounted the
twain and drew up for battle fain; and Sharrkan was going forth to
the plain, but behold, more than one half of the Franks dismounted
and remained on foot before one of them who was mounted, till
they reached the midst of the battle-plain. Sharrkan looked at that
horseman and lo! he was their chief. He was clad in a surcoat of blue
satin and a close-ringed mail shirt; his face was as the moon when it
rises and no hair was upon his cheeks. He hent in hand an Indian
scymitar and he rode a sable steed with a white blaze on brow, like
a dirham; and he smote the horse with heel till he stood almost in
the midst of the field when, signing to the Moslems, he cried out in
fluent Arab speech, "Ho, Sharrkan! Ho, son of Omar bin al-Nu'uman!
Ho, thou who forcest fortalice and overthrowest cities and countries!
up and out to battle-bout, and blade single-handed wield with one](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-55-320.jpg)
![by a woman."[216]
When Sharrkan heard this, he raised his eyes to
the Frank's face and gazing steadfastly at him, recognized in him
Princess Abrizah with whom that pleasant adventure had befallen
him in the convent; whereupon he cast brand from hand and, kissing
the earth before her, asked her, "What moved thee to a deed like
this?"; and she answered, "I desired to prove thy prowess afield and
test thy doughtiness in tilting and jousting. These that are with me
are my handmaids, and they are all clean maids; yet they have
vanquished thy horsemen in fair press and stress of plain; and had
not my steed stumbled with me, thou shouldst have seen my might
and prowess in combat." Sharrkan smiled at her speech and said,
"Praise be to Allah for safety and for my reunion with thee, O Queen
of the age." Then she cried out to her damsels to loose the twenty
captives of Sharrkan's troop and dismount. They did as she bade
and came and kissed the earth before her and Sharrkan who said to
them, "It is the like of you that Kings keep in store for the need-
hour." Then he signed to his comrades to salute the Princess; so all
alighted and kissed the earth before her, for they knew the story.
After this, the whole two hundred took horse, and fared on night
and day for six days' space, till they drew near to Baghdad, when
they halted and Sharrkan bade Abrizah and her handmaids doff the
Frankish garb that was on them——And Shahrazad perceived the
dawn of day and ceased saying her permitted say.
Now when it was the Fifty-first Night,
She said, It hath reached me, O auspicious King, that Sharrkan bade
Princess Abrizah and her damsels doff the garb that was on them
and don the garments of daughters of Greece; and thus did they.
Then he despatched a company of his companions to Baghdad to
acquaint his father Omar bin al-Nu'uman, with his arrival and report
that he was accompanied by Princess Abrizah, daughter of King
Hardub, Lord of Græcia-land. They halted forthright in the place they
had reached, and Sharrkan also halted and all nighted there; and](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-57-320.jpg)
![when Almighty Allah made morning dawn, Sharrkan and his
company and Abrizah and her company took horse and fared on
towards the city; when lo! on the way they met the Wazir Dandan,
who had come out amongst a thousand horse, to honour Abrizah
and Sharrkan, by especial commandment of King Omar Son of Al-
Nu'uman. When the two drew near, they turned towards them and
kissed ground before them; then they mounted again and escorted
them into the city and went up with them to the palace. Sharrkan
walked in to his father, who rose and embraced him and questioned
him of his case. So he told him all that Abrizah had told him, and
what had passed between them and said, "She hath parted from her
sire and departed from her reign and hath chosen to take part with
us and to make her abode with us; and indeed (he said to his
father) the King of Constantinople hath plotted to do us a mischief,
because of his daughter Sophia, for that the King of Greece had
made known to him her story and the cause of her being given to
thee; and he (the Grecian King) not knowing her to be daughter of
King Afridun, Lord of Constantinople; and, had he known that, he
would not have bestowed her upon thee, but he would have
restored her to her parent. And of a verity (he continued) we were
saved from these perils only by the Lady Abrizah, and never saw we
a more valiant than she." And he went on to tell his father all that
had passed from first to last of the wrestling and the single-fighting.
When King Omar heard the story of Sharrkan, Abrizah was exalted in
his eyes, and he longed to see her and question her. Thereupon
Sharrkan went out to her and said, "The King calleth for thee;" she
replied, "I hear and I obey;" and he took her and brought her in to
his father, who was seated on his throne and who, having dismissed
his high officers, was attended only by his eunuchs. The Princess
entered and kissing the ground between his hands, saluted him in
choice terms. He was amazed at her eloquent speech and thanked
her for her dealing with his son Sharrkan and bade her be seated.
So she sat down and unveiled her face;[217]
and, when the King saw
her beauty, his reason fled his head and he made her draw near and
showed her favour, appointing her an especial palace for herself and
her damsels, and assigning them solde and allowances. Then began](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-58-320.jpg)
![standing with thee, overwhelmed as I am with thy favours and thy
benefits? And, furthermore, Allah hath blessed me with two children
by thee, a son and a daughter." Her reply pleased the King and after
leaving her, he set apart for her and her children a wondrous fine
palace. Moreover, he appointed for them eunuchs and attendants
and doctors of law and doctors of philosophy and astrologers and
physicians and surgeons to do them service; and in every way he
redoubled his favour and entreated them with the best of treatment.
And presently he returned to the palace of his dominion and to his
Court where he distributed justice among the lieges. So far
concerning him and Sophia and her children; but in the matter of
Abrizah the King was greatly occupied with love of her and burnt
with desire of her night and day; and every night, he would go in to
her and converse with her and pay his court to her, but she gave
him no answer, only saying, "O King of the age! I have no desire for
men at this present." When he saw her withdraw from him, his
passion waxed hotter and his longing and pining increased until,
when weary of this, he summoned his Wazir Dandan and, opening
his very heart to him, told him of his love for Princess Abrizah,
daughter of Hardub, and informed him how she refused to yield to
his wishes and how desire for her was doing him to die, for that he
could get no grace of her. The Wazir, hearing these words, said to
the King, "As soon as it is dark night, take thou a piece of Bhang the
measure of a miskal, about an ounce, and go in to her and drink
somewhat of wine with her. When the hour of ending the carousal
shall draw near, fill her a last cup and dropping therein the Bhang,
give it to her to drink, and she will not reach her sleeping-chamber
ere the drug take effect on her. Then do thou go in to her and take
thy will of her; and such is my advice."[218]
"Thy rede is aright,"
quoth the King, and seeking his treasury, he took thence a piece of
concentrated Bhang, if an elephant smelt it he would sleep from
year to year. This he put in his bosom-pocket and waited till some
little of the night went by, when he betook himself to the palace of
Princess Abrizah, who seeing him stood up to receive him; but he
bade her sit down. So she sat down, and he sat by her, and he
began to talk with her of wine and wassail, whereupon she furnished](https://image.slidesharecdn.com/21057-250316130751-e2a13427/85/Immunoelectron-Microscopy-Methods-and-Protocols-1st-Edition-David-M-Rancour-61-320.jpg)
Immunoelectron Microscopy Methods and Protocols 1st Edition David M. Rancour
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- 5. Immunoelectron Microscopy Methods and Protocols 1st Edition David M. Rancour Digital Instant Download Author(s): David M. Rancour, Steven K. Backues, Sebastian Y. Bednarek (auth.), Steven D. Schwartzbach, Tetsuaki Osafune (eds.) ISBN(s): 9781607617839, 1607617838 Edition: 1 File Details: PDF, 13.48 MB Year: 2010 Language: english
- 7. METHODS IN MOLECULAR BIOLOGY TM Series Editor John M. Walker School of Life Sciences University of Hertfordshire Hatfield, Hertfordshire, AL10 9AB, UK For other titles published in this series, go to www.springer.com/series/7651
- 9. Immunoelectron Microscopy Methods and Protocols Edited by Steven D. Schwartzbach DepartmentofBiology,UniversityofMemphis,Memphis,TN,USA Tetsuaki Osafune DepartmentofLifeScience,NipponSportScienceUniversity,Yokohama,Japan
- 10. Editors Steven D. Schwartzbach Department of Biology University of Memphis 3774 Walker Avenue Memphis, TN 38152 USA sdschwrt@memphis.edu Tetsuaki Osafune Department of Life Science Nippon Sport Science University Kamosida 1221-1 227-0033 Yokohama Japan osafunet@aol.com ISSN 1064-3745 e-ISSN 1940-6029 ISBN 978-1-60761-782-2 e-ISBN 978-1-60761-783-9 DOI 10.1007/978-1-60761-783-9 Springer New York Dordrecht Heidelberg London Library of Congress Control Number: 2010929610 © Springer Science+Business Media, LLC 2010 All rights reserved. This work may not be translated or copied in whole or in part without the written permission of the publisher (Humana Press, c/o Springer Science+Business Media, LLC, 233 Spring Street, New York, NY 10013, USA), except for brief excerpts in connection with reviews or scholarly analysis. Use in connection with any form of information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed is forbidden. The use in this publication of trade names, trademarks, service marks, and similar terms, even if they are not identified as such, is not to be taken as an expression of opinion as to whether or not they are subject to proprietary rights. Cover illustration: A 3D solid surface rendering showing that the immunogold-labeled small subunit of ribulose-bis- phosphate carboxylase, yellow dots, is concentrated in the propyrenoid, blue, rather than being uniformly distributed throughout the prolamellar body, red, and proplastid. The 3D reconstruction of the distribution of ribulose-bis- phosphate carboxylase is shown in the left, while the 3D reconstruction of the propyrenoid and prolamellar body superimposed upon the 3D distribution of ribulose-bis-phosphate carboxylase is shown in the right. Printed on acid-free paper Humana Press is part of Springer Science+Business Media (www.springer.com)
- 11. Preface Cell biology is the science of correlating cell structure and function. The electron micro- scopist obtains high-resolution pictures of the intricate structures found in cells. Electron tomography and serial sections can be used to reconstruct the three-dimensional structure of the cell and its organelles. Pictures are worth a thousand words, but they are unable to provide information regarding the function of the intricate structures found in cells and their macromolecular composition. The biochemist and molecular biologist determine the functions of the molecules, macromolecular complexes, and organelles found within cells. They isolate individual cellular constituents and reconstruct vital cellular processes. These in vitro experiments provide a detailed understanding of cellular function. Organelle isolation provides a method to place macromolecular functions within a structural context. Understanding structure–function relationships through organelle isolation has restricted utility because organelles cannot be isolated from every organism, not every organelle can be isolated free of contamination by other organelles, suborganellular compartments often cannot be purified for biochemical characterization, and, when a protein is recovered in mul- tiple organelles, it is often difficult to distinguish true localization from contamination artifacts. Immunoelectron microscopy is the technique that bridges the information gap between biochemistry, molecular biology, and ultrastructural studies placing macromolec- ular functions within a cellular context. Immunoelectron microscopy can be used on vir- tually every unicellular and multicellular organism. The only requirements are suitable fixation protocols and the availability of an antibody to the molecule whose structural location is to be determined. Structure–function relationships can be determined even when it is impossible to purify the organelle or suborganellular compartment containing the macromolecules being studied. Most importantly, immunoelectron microscopy is a totally objective procedure that is not dependent on conjectures as to where the protein is localized and thus which organelles to isolate for biochemical studies. Two examples from our own work demonstrate how immunoelectron microscopy provides unexpected insights into structure–function relationships. Immunoelectron microscopy first demonstrated that the Euglena light harvesting chlorophyll a/b binding protein of photosystem II (LHCPII) was found in the Golgi apparatus prior to its pres- ence in the chloroplast. This finding was the impetus for detailed biochemical studies that elucidated a new mechanism for chloroplast protein import: transport from the ER to the Golgi apparatus to the chloroplast. Immunoelectron microscopy identified the pyrenoid as the site of the enzyme ribulose 1–5 bisphosphate carboxylase/oxygenase (RUBISCO) showing that the enzyme moves from the stroma to the pyrenoid at cell cycle phases when enzyme activity is high while the pyrenoid disappears and RUBISCO redistributes back to the stroma at cell cycle phases when enzyme activity is low. Biochemical studies identified the activity changes but organelle fractionation experiments never identified this change in suborganellular localization because pyrenoids could not be isolated. At all cell cycle stages, the enzyme was recovered in the soluble chloroplast fraction. v
- 12. vi Preface The successful application of immunoelectron microscopy requires combining the tools of the molecular biologist with those of the microscopist. From the molecular biology toolbox, this volume will present methods for antigen production by protein expression in bacterial cells and by expression of epitope tagged proteins in plant and animal cells. Methods for production of anti-peptide, monoclonal, and polyclonal anti- bodies will be presented. From the microscopy toolbox, this volume will present methods for cryoultramicrotomy and rapid freeze-replacement fixation which have the advantage of retaining protein antigenicity at the expense of ultrastructural integrity as well as chemical fixation methods that maintain structural integrity while sacrificing protein antigenicity. Plants and algae contain cell walls, vacuoles, and other structures which present barriers to antibody penetration and complicate fixation. Due to these problems, separate chap- ters will discuss fixation and immunolabeling protocols for animals, plants, and algae. Pre- and post-embedding immunogold labeling protocols will be presented. Pre-embedding methods perform immunogold labeling before ultrathin sections are prepared from resin- embedded samples resulting in greater sensitivity and better microstructure preservation. Post-embedding methods perform immunolabeling after ultrathin sections are prepared from resin-embedded samples resulting in decreased antigenicity. The detailed methods and notes will facilitate choosing the best method for the antibody and biological material to be studied. Finally, methods will be presented for immunogold labeling of two anti- gens for protein colocalization studies, for three-dimensional reconstruction of intracellu- lar antigen distribution, for immunogold labeling of DNA, and for immunogold scanning electron microscopy. It is our hope that the toolbox created by this volume will facilitate an increased understanding of structure–function relationships. Steven D. Schwartzbach Tetsuaki Osafune
- 13. Contents Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v Contributors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ix PART I MOLECULAR TOOLBOX 1. Protein Antigen Expression in Escherichia coli for Antibody Production . . . . . 3 David M. Rancour, Steven K. Backues, and Sebastian Y. Bednarek 2. Expression of Epitope-Tagged Proteins in Plants . . . . . . . . . . . . . . . . . 21 Takuya Furuichi 3. Expression of Epitope-Tagged Proteins in Arabidopsis Leaf Mesophyll Protoplasts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33 Young-Hee Cho and Sang-Dong Yoo 4. Transient Expression of Epitope-Tagged Proteins in Mammalian Cells . . . . . . 43 Melanie L. Styers, Jason Lowery, and Elizabeth Sztul 5. Production and Purification of Polyclonal Antibodies . . . . . . . . . . . . . . . 63 Masami Nakazawa, Mari Mukumoto, and Kazutaka Miyatake 6. Production and Purification of Monoclonal Antibodies . . . . . . . . . . . . . . 75 Masami Nakazawa, Mari Mukumoto, and Kazutaka Miyatake 7. Production of Antipeptide Antibodies . . . . . . . . . . . . . . . . . . . . . . . 93 Bao-Shiang Lee, Jin-Sheng Huang, G.D. Lasanthi P. Jayathilaka, Syed S. Lateef, and Shalini Gupta 8. Preparation of Colloidal Gold Particles and Conjugation to Protein A, IgG, F(ab’)2, and Streptavidin . . . . . . . . . . . . . . . . . . . . . . . . . . 109 Sadaki Yokota PART II MICROSCOPY TOOLBOX 9. Immunoelectron Microscopy of Chemically Fixed Developing Plant Embryos . . 123 Tetsuaki Osafune and Steven D. Schwartzbach 10. Pre-embedding Immunogold Localization of Antigens in Mammalian Brain Slices . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 133 Thomas Schikorski 11. Pre-embedding Immunoelectron Microscopy of Chemically Fixed Mammalian Tissue Culture Cells . . . . . . . . . . . . . . . . . . . . . . . . . 145 Haruo Hagiwara, Takeo Aoki, Takeshi Suzuki, and Kuniaki Takata 12. Immunoelectron Microscopy of Cryofixed and Freeze-Substituted Plant Tissues . . . . . . . . . . . . . . . . . . . . . . . 155 Miyuki Takeuchi, Keiji Takabe, and Yoshinobu Mineyuki vii
- 14. viii Contents 13. In Vivo Cryotechniques for Preparation of Animal Tissues for Immunoelectron Microscopy . . . . . . . . . . . . . . . . . . . . . . . . . 167 Shinichi Ohno, Nobuhiko Ohno, Nobuo Terada, Sei Saitoh, Yurika Saitoh, and Yasuhisa Fujii 14. Immunoelectron Microscopy of Cryofixed Freeze-Substituted Mammalian Tissue Culture Cells . . . . . . . . . . . . . . . . . . . . . . . . . 181 Akira Sawaguchi 15. Immunoelectron Microscopy of Cryofixed Freeze-Substituted Saccharomyces cerevisiae . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191 Jindriska Fiserova and Martin W. Goldberg 16. High-Resolution Molecular Localization by Freeze-Fracture Replica Labeling . . 205 Akikazu Fujita and Toyoshi Fujimoto 17. Pre-embedding Electron Microscopy Methods for Glycan Localization in Chemically Fixed Mammalian Tissue Using Horseradish Peroxidase-Conjugated Lectin . . . . . . . . . . . . . . . . . . . . . . . . . . 217 Yoshihiro Akimoto and Hayato Kawakami 18. Pre-embedding Nanogold Silver and Gold Intensification . . . . . . . . . . . . 225 Akitsugu Yamamoto and Ryuichi Masaki 19. The Post-embedding Method for Immunoelectron Microscopy of Mammalian Tissues: A Standardized Procedure Based on Heat-Induced Antigen Retrieval . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 237 Shuji Yamashita 20. Double-Label Immunoelectron Microscopy for Studying the Colocalization of Proteins in Cultured Cells . . . . . . . . . . . . . . . . . 249 Haruo Hagiwara, Takeo Aoki, Takeshi Suzuki, and Kuniaki Takata 21. Serial Section Immunoelectron Microscopy of Algal Cells . . . . . . . . . . . . 259 Tetsuaki Osafune and Steven D. Schwartzbach 22. Freeze-Etch Electron Tomography for the Plasma Membrane Interface . . . . . 275 Nobuhiro Morone 23. Localization of rDNA at Nucleolar Structural Components by Immunoelectron Microscopy . . . . . . . . . . . . . . . . . . . . . . . . . 287 Seiichi Sato and Yasushi Sato 24. Immunogold Labelling for Scanning Electron Microscopy . . . . . . . . . . . . 297 Martin W. Goldberg and Jindriska Fiserova 25. Horseradish Peroxidase as a Reporter Gene and as a Cell-Organelle- Specific Marker in Correlative Light-Electron Microscopy . . . . . . . . . . . . 315 Thomas Schikorski 26. Monitoring Rapid Endocytosis in the Electron Microscope via Photoconversion of Vesicles Fluorescently Labeled with FM1-43 . . . . . . . . . 329 Thomas Schikorski Subject Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 347 .
- 15. Contributors YOSHIHIRO AKIMOTO • Department of Anatomy, Kyorin University School of Medicine, Mitaka, Tokyo, Japan TAKEO AOKI • Department of Anatomy and Cell Biology, Gunma University Graduate School of Medicine, Gunma, Japan STEVEN K. BACKUES • Department of Biochemistry, University of Wisconsin, Madison, WI, USA SEBASTIAN Y. BEDNAREK • Department of Biochemistry and Great Lakes Bioenergy Research Center, University of Wisconsin, Madison, WI, USA YOUNG-HEE CHO • Department of Molecular Biology, Massachusetts General Hospital, Boston, MA, USA; Laboratory of Plant Signaling Systems Biology, Department of Biolog- ical Science, College of Natural Sciences, Sungkyunkwan University, Suwon, Korea JINDRISKA FISEROVA • School of Biological and Biomedical Sciences, Durham University, Durham, UK YASUHISA FUJII • Department of Anatomy and Molecular Histology, Interdisciplinary Graduate School of Medicine and Engineering, University of Yamanashi, Yamanashi, Japan TOYOSHI FUJIMOTO • Department of Anatomy and Molecular Cell Biology, Nagoya Uni- versity Graduate School of Medicine, Nagoya, Japan AKIKAZU FUJITA • Department of Anatomy and Molecular Cell Biology, Nagoya Univer- sity Graduate School of Medicine, Nagoya, Japan TAKUYA FURUICHI • Research Institute for Bioresources, Okayama University, Okayama, Japan MARTIN W. GOLDBERG • School of Biological and Biomedical Sciences, Durham Univer- sity, Durham, UK SHALINI GUPTA • Protein Research Laboratory, Research Resources Center, University of Illinois at Chicago, Chicago, IL, USA HARUO HAGIWARA • Department of Occupational Therapy, School of Health Sciences, Faculty of Medicine, Gunma University, Gunma, Japan JIN-SHENG HUANG • Protein Research Laboratory, Research Resources Center, University of Illinois at Chicago, Chicago, IL, USA G.D. LASANTHI P. JAYATHILAKA • Protein Research Laboratory, Research Resources Cen- ter, University of Illinois at Chicago, Chicago, IL, USA HAYATO KAWAKAMI • Department of Anatomy, Kyorin University School of Medicine, Mitaka, Tokyo, Japan SYED S. LATEEF • Protein Research Laboratory, Research Resources Center, University of Illinois at Chicago, Chicago, IL, USA BAO-SHIANG LEE • Protein Research Laboratory, Research Resources Center, University of Illinois at Chicago, Chicago, IL, USA JASON LOWERY • Department of Cell Biology, University of Alabama at Birmingham, Birmingham, AL, USA ix
- 16. x Contributors RYUICHI MASAKI • Department of Physiology, Kansai Medical University, Moriguchi, Osaka, Japan YOSHINOBU MINEYUKI • Department of Life Science, Graduate School of Life Science, University of Hyogo, Hyogo, Japan KAZUTAKA MIYATAKE • Department of Applied Biological Chemistry, Osaka Prefecture University, Osaka, Japan NOBUHIRO MORONE • National Institute of Neuroscience, National Center of Neurology and Psychiatry, Tokyo, Japan; Institute for Integrated Cell-Material Sciences (iCeMS), Kyoto University, Kyoto, Japan MARI MUKUMOTO • Department of Applied Biological Chemistry, Osaka Prefecture Uni- versity, Osaka, Japan MASAMI NAKAZAWA • Department of Applied Biological Chemistry, Osaka Prefecture University, Osaka, Japan NOBUHIKO OHNO • Department of Anatomy and Molecular Histology, Interdisciplinary Graduate School of Medicine and Engineering, University of Yamanashi, Yamanashi, Japan SHINICHI OHNO • Department of Anatomy and Molecular Histology, Interdisciplinary Graduate School of Medicine and Engineering, University of Yamanashi, Yamanashi, Japan TETSUAKI OSAFUNE • Department of Life Science, Nippon Sport Science University, Yoko- hama, Japan DAVID M. RANCOUR • Department of Biochemistry and Great Lakes Bioenergy Research Center, University of Wisconsin, Madison, WI, USA SEIICHI SATO • Department of Biology, Faculty of Science, Ehime University, Matsuyama, Japan YASUSHI SATO • Department of Biology, Faculty of Science, Ehime University, Matsuyama, Japan SEI SAITOH • Department of Anatomy and Molecular Histology, Interdisciplinary Grad- uate School of Medicine and Engineering, University of Yamanashi, Yamanashi, Japan YURIKA SAITOH • Department of Anatomy and Molecular Histology, Interdisciplinary Graduate School of Medicine and Engineering, University of Yamanashi, Yamanashi, Japan AKIRA SAWAGUCHI • Department of Anatomy, Ultrastructural Cell Biology, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan THOMAS SCHIKORSKI • Neuroscience Department, Universidad Central del Caribe, Bayamon, Puerto Rico, USA STEVEN D. SCHWARTZBACH • Department of Biology, University of Memphis, Memphis, TN, USA MELANIE L. STYERS • Department of Biology, Chemistry, and Mathematics, University of Montevallo, Montevallo, AL, USA TAKESHI SUZUKI • Department of Anatomy and Cell Biology, Gunma University Grad- uate School of Medicine, Gunma, Japan ELIZABETH SZTUL • Department of Cell Biology, University of Alabama at Birmingham, Birmingham, AL, USA KEIJI TAKABE • Division of Forest and Biomaterials Science, Graduate School of Agricul- ture, Kyoto University, Kyoto, Japan KUNIAKI TAKATA • Department of Anatomy and Cell Biology, Gunma University Grad- uate School of Medicine, Gunma, Japan
- 17. Contributors xi MIYUKI TAKEUCHI • Department of Life Science, Graduate School of Life Science, Uni- versity of Hyogo, Hyogo, Japan NOBUO TERADA • Department of Anatomy and Molecular Histology, Interdisciplinary Graduate School of Medicine and Engineering, University of Yamanashi, Yamanashi, Japan AKITSUGU YAMAMOTO • Department of Animal Bio-Science, Faculty of Bio-Science, Nagahama Institute of Bio-Science and Technology, Nagahama, Japan SHUJI YAMASHITA • Electron Microscope Laboratory, School of Medicine, Keio University, Tokyo, Japan SADAKI YOKOTA • Section of Functional Morphology, Faculty of Pharmaceutical Science, Nagasaki International University, Nagasaki, Japan SANG-DONG YOO • Laboratory of Plant Signaling Systems Biology, Department of Biolog- ical Science, College of Natural Sciences, Sungkyunkwan University, Suwon, Korea
- 19. Chapter 1 Protein Antigen Expression in Escherichia coli for Antibody Production David M. Rancour, Steven K. Backues, and Sebastian Y. Bednarek Abstract Escherichia coli is a frequently used expression system for the generation of protein encoded by genes from diverse kingdoms and, thus, it is well suited for the production of protein antigens for antibody generation. It is a system of choice for many due to factors such as (1) the commercial availability of a vast array of reagents and materials needed for cloning, expression, and purification and (2) the potential high protein yields that can be acquired in a timely and cost-effective manner. This chapter will focus on (1) the general principles to keep in mind when choosing an antigen to express and (2) the use of a modified pGEX vector system (Rancour et al., J. Biol. Chem. 279:54264–54274, 2004) to use in its expression. Simplified protocols are provided for (1) assessing the expression of your protein, (2) testing whether your protein is or is not expressed as a soluble product, (3) performing bulk purifications of soluble or insoluble E. coli-expressed protein to acquire enough to be used for a complete immunization protocol, and (4) an optional procedure for epitope tag removal from your expressed protein of interest in order to avoid the unnecessary and sometimes unwanted production of antibodies against the fusion protein affinity chromatography tag. These four procedures have been used extensively and successfully in our lab as a basis for the production of recombinant protein and subsequent antibody production. Key words: Escherichia coli, protein, expression, purification, glutathione-S-transferase, GST, epitope tag, antigen, tobacco etch virus (TEV) protease. 1. Introduction Protein expression in Escherichia coli is a frequently used tool for the generation of protein encoded by genes from diverse king- doms and, thus, it is well suited for the production of protein anti- gens for antibody generation. To facilitate the generation of useful antibodies against a target protein, there are several characteristics of the protein that need to be assessed prior to commencing with S.D. Schwartzbach, T. Osafune (eds.), Immunoelectron Microscopy, Methods in Molecular Biology 657, DOI 10.1007/978-1-60761-783-9_1, © Springer Science+Business Media, LLC 2010 3
- 20. 4 Rancour, Backues, and Bednarek expression. Taking the following details into consideration while determining the best antigen to express will greatly increase the chance that quality antibodies will be generated. The first consideration is whether the protein of interest is known or predicted to be a soluble, a peripheral membrane, or an integral membrane protein. Soluble proteins or domains are much easier to work with in terms of expression, purification, and, in some case, immunization protocols. For peripheral and integral membrane proteins, select cDNA fragments predicted to encode hydrophilic soluble domains of the protein of inter- est (i.e., domains lacking long contiguous hydrophobic amino acid stretches or transmembrane segments as predicted by hydro- pathicity analysis using the Kyte–Doolittle (1) and/or Hopp– Woods (2) algorithms or utilizing newer methods of soluble pro- tein prediction such as the method of Smialowski et al. (3)). Insoluble proteins can also be expressed and enriched as inclusion bodies in E. coli and then subsequently used for immunization. However, soluble proteins or protein domains most likely best represent the native folding organization of the protein in its cel- lular context and thus would be the optimal antigen to produce antibodies that would be used for immunocytochemistry. Addi- tional drawbacks to using insoluble proteins include the purity of the protein not being high enough to generate specific anti-sera against the antigen, and affinity purification of specific antibod- ies becomes problematic (but not impossible) using the original antigen. Insoluble proteins can limit the use of standard liquid chromatography purification methods sans attempts at denatu- ration and renaturation procedures. Therefore, soluble proteins or domains should be the first choice for easy antigen produc- tion and purification to ensure a clean antigen to induce a specific immunological response. The second consideration is whether the protein of interest is post-translationally modified. If so, does this modification influ- ence its functionality and/or localization? Proteins from eukary- otes are commonly post-translationally modified. For example, in cases where these proteins are extensively decorated with carbohy- drates, these modifications may adversely influence the reactivity of antibodies raised against an unmodified protein expressed in E. coli. Conversely, if you are interested in antibodies against anti- gens with specific modifications, you will need to keep in mind that most eukaryotic cell modifications do not take place in E. coli and thus you will need to seek other methods for antigen production. The third consideration is whether the protein of interest is either a homo- or hetero-oligomer. Oligomerization may influ- ence the ability to express the protein of interest as a soluble pro- tein. A self-assembling soluble homo-oligomer [i.e., AtCDC48A (4)] is much easier to produce than a subunit of a multimeric,
- 21. Protein Antigen Expression in E. coli for Antibody Production 5 membrane-associated complex that needs specific chaperones for assembly. In the latter case, choosing a soluble domain fragment of the protein may be a better antigen choice. The fourth consideration is whether the antigen is a prod- uct of a conserved gene family or relatively unique. To gener- ate isoform-specific antibodies, expression of sequence divergent domains will be required. In cases where the divergent amino acid sequence is limited to short stretches (12–15 aa), synthetic pep- tides coupled to a carrier protein may be a better antigen choice. Alternatively, choosing a fragment of the protein that does not contain a highly conserve protein domain (e.g., Walker ATPase) could aid in minimizing cross-reactivity of your anti-serum. Protein expression vectors for E. coli typically differ in the pro- moter/repressor system used for gene expression regulation and the type/position of epitope tags translationally fused to the pro- tein product and are available from various commercial and aca- demic sources. A modified pGEX4T plasmid expression system (4) encoding N-terminally fused glutathione-S-transferase (∼26- kDa soluble protein from Schistosoma japonicum) is used in these protocols. The GST epitope tag has been shown to aid in increas- ing protein solubility, facilitating purification (5, 6), and allowing for higher yields (7). The RosettaTM (DE3) pLysS E. coli strain (Novagen-EMD Biosciences) is used for protein expression in these protocols because the strain is protease deficient and con- tains a plasmid encoding six tRNAs underutilized by E. coli to alleviate codon bias which sometimes reduces protein expression of genes from divergent organismal origins. The epitope tag is normally removed from an antigen to min- imize the incidental production of antibodies to the tag in addi- tion to the desired antigen. Removal of the epitope tag prior to use as an immunogen also facilitates subsequent affinity purifica- tion of antibodies to the protein of interest by allowing the origi- nal fusion protein to be used for affinity purification. A very use- ful feature of the modified pGEX4T-TEV vector system (4) used in these protocols is the inclusion of both the original thrombin and an added tobacco etch virus (TEV) sequence-specific cleavage sites between the GST tag and the protein insert. The specificity of TEV for its recognition sequence is more stringent than that of thrombin (8), thereby reducing unwanted cleavage of your pro- tein of interest. In addition to its utility for tag removal, the use of TEV is quite cost effective because it can be easily produced in-house (9) from publicly available expression strains (10). TEV may also be purchased from several commercial sources including Promega, Eton Biosciences, and Invitrogen. After the gene encoding the antigen is cloned into the mod- ified pGEX4T-TEV vector, it should be first be transformed into the RosettaTM (DE3) pLysS or other protein expression compati- ble E. coli strains. To produce a purified tag-free soluble bacterially
- 22. 6 Rancour, Backues, and Bednarek expressed protein for use as an immunogen, the resulting clones must first be tested via small-scale induction to verify expression, and then subsequently by small-scale fractionation to determine the solubility of the protein product. Once the expression and solubility of the protein has been verified, large-scale expression and purification of the soluble protein by affinity chromatography using glutathione resin can be initiated. GST tag removal from purified soluble fusion proteins using sequence-specific proteases followed by protease removal by affinity chromatography is the final step in preparing a purified soluble bacterially expressed pro- tein for use as an immunogen. For insoluble proteins, inclusion bodies are isolated and used for immunization. In this chapter, we present protocols for the preparation of tag-free soluble and insoluble bacterially expressed proteins for use as immunogens. 2. Materials 2.1. SDS-PAGE (Sodium Dodecylsulfate- Polyacrylamide Gel Electrophoresis) 1. Precast Ready Gel Tris–HCl SDS-PAGE gels, 4–15% acry- lamide, Tris/glycine/SDS buffer (Bio-Rad Laboratories, Inc., Hercules, CA). 2. Small vertical electrophoresis unit (Hoefer, Inc., San Fran- cisco, CA). 3. Power Supply (Bio-Rad Laboratories, Inc., Hercules, CA). 4. 10× SDS-PAGE running buffer: Mix 30.3 g Tris base, 144 g glycine, 10 g SDS, and distilled water (dH2O) to a final volume of 1 L; dilute with dH2O to 1× strength for use. 5. 5× SDS-PAGE sample buffer (5× SSB): Mix 3.9 mL 2 M Tris–HCl pH 6.8, 2.5 g SDS, 12.5 mL glycerol, 8 mg bro- mophenol blue, 6.25 mL β-mercaptoethanol, and double distilled water (ddH2O) to a final volume of 25 mL. Use at 2× strength (2× SSB): 4 mL 5× SSB diluted with ddH2O to final volume of 10 mL. 6. Protein Molecular Weight Markers: SDS-PAGE broad range standards (Bio-Rad Laboratories, Inc., Hercules, CA) diluted 1:40 in 2× SSB. 7. Coomassie stain for protein gels: 0.1% (w/v) Coomassie R- 250 dissolved in fixative [40% (v/v) methanol, 10% (v/v) glacial acetic acid]; store capped at room temperature. 8. Coomassie destain solution for protein gels: 40% (v/v) methanol, 10% (v/v) glacial acetic acid; store capped at room temperature. 9. Kimwipes (Kimberly-Clark Corporation, Irving, TX). 10. Gel drying kit (Promega Corp., Madison, WI).
- 23. Protein Antigen Expression in E. coli for Antibody Production 7 2.2. Analytical Scale Test of GST-Fusion Protein Expression 1. The modified pGEX4T plasmid engineered with a TEV protease cleavage site between endogenous Thrombin cleavage site and MCS (see Note 1) or pGEX4T plasmid (GE Healthcare Bio-Sciences Corp., Piscataway, NJ) containing the coding sequence for the protein/ domain of interest in RosettaTM (DE3) pLysS E. coli protein expression strain (Novagen-EMD Biosciences, Madison, WI). 2. LB broth: Mix 10.0 g Tryptone (BD Biosciences, San Jose, CA), 5.0 g yeast extract (BD Biosciences, San Jose, CA), 10.0 g sodium chloride, and ddH2O to a final volume of 1 L; autoclave 20 min at 121◦C; cool to 55◦C before adding antibiotics. 3. Solid LB plates: Add 15.0 g/L Bactoagar (BD Biosciences, San Jose, CA) to freshly prepared LB broth prior to auto- claving. After autoclaving, cool broth to 55◦C in a water bath, add antibiotic, mix by swirling, and aseptically pour a thin layer into 100-mm × 15-mm sterile plastic Petri dishes. Allow plates to cool to RT and store plates inverted in a plastic sleeve at 4◦C. 4. Antibiotics: Carbenicillin (1,000×): 50.0 mg/mL in 50% (v/v) ethanol. Chloramphenicol (1,000×): 34.0 mg/mL in 95% (v/v) ethanol. Store both at −20◦C. 5. Isopropyl-β-D-thiogalactopyranoside (IPTG) (Sigma- Aldrich, St. Louis, MO): 500 mM stock solution in ddH2O, sterile filter, and store at −20◦C. 6. Tris-buffered saline pH 7.4 (TBS pH 7.4; 10× strength): Mix 80.0 g NaCl, 0.2 g KCl, 30.0 g Tris base, and 800 mL of ddH2O. With concentrated HCl, adjust pH to 7.4 and then bring to a final volume of 1 L with ddH2O. Use at 1× strength and store at room temperature. 7. Spectrophotometer (600 nm capability) (GE Healthcare Bio-Sciences Corp., Piscataway, NJ). 8. Micro-tip sonicator (Branson Ultrasonics, Danbury, CT). 2.3. Protein Expression Test for Solubility and Capacity to Bind Glutathione– Sepharose Items in Section 2.2 are also included: 1. β-Mercaptoethanol (β-ME) (Sigma-Aldrich, St. Louis, MO). 2. NP-40 (Calbiochem-EMD Biosciences, San Diego, CA); stock solution of 20% (v/v) in dH2O; store at room tem- perature. 3. Glutathione–SepharoseTM 4 Fast-Flow (GE Life- Sciences, Inc., Piscataway, NJ); working stock of 25% (v/v) in 1× TBS pH 7.4, supplemented with 0.1% (v/v) NP-40 and 0.02% (w/v) sodium azide; store at 4◦C.
- 24. 8 Rancour, Backues, and Bednarek 2.4. Large-Scale Expression and Purification Items in Section 2.3 are also included: 1. Phenylmethanesulfonyl fluoride (PMSF) (Sigma-Aldrich, St. Louis, MO) 200 mM stock in dry isopropanol (see Note 2). 2. Poly-Prep Chromatography Column (Bio-Rad Laborato- ries, Inc., Hercules, CA). 3. Adenosine 5’-triphosphate, disodium salt (Sigma-Aldrich, St. Louis, MO) made into 100 mM stock solution with dH2O, stored at −20◦C in small (∼200 μL) aliquots to avoid freeze-thaw cycles. 4. MgCl2: 1 M stock solution in dH2O, autoclave, and store at room temperature. 5. L-Glutathione, reduced (Sigma-Aldrich, St. Louis, MO). 6. Tris-buffered saline pH 8.0 (TBS pH 8.0; 10× strength): Mix 8.0 g NaCl, 0.02 g KCl, 3.0 g Tris base, and dH2O for a final volume of 100 mL adjusting the pH to 8.0 with concentrated HCl; use at 1× strength, store at room tem- perature. 7. Elution solution: 1× TBS pH 8.0 + 15 mM L- glutathione (reduced) (46 mg/10.0 mL). Prepare imme- diately before use. 8. Pierce 660 nm Protein Assay Kit (Thermo Scientific Pierce Protein Research Products, Rockford, IL). 9. Microplate spectrophotometer (660 nm capability) (Bio- Tek Instruments, Winooski, VT). 10. 96-Well, flat bottom microtiter plates (Greiner Bio-One North America, Monroe, NC). 11. Dialysis tubing (MWCO 12–14,000, 1 cm flat width, 0.3 mL/cm; Spectra/Por; Spectrum Laboratories, Inc., Ran- cho Dominguez, CA) and clips. 12. Glycerol. 13. Buffer A (need 50 mL): 25 mM HEPES–NaOH pH 7.5, 1 mM EDTA-NaOH pH 7.5. 14. Lysis buffer: To 50.0 mL buffer A add 50.0 μL NP- 40 (100% stock), 5.0 mg lysozyme, 0.5 mg RNAse- A, 2.5 mg DNAse I, 100.0 μL MgCl2 (1 M stock). 15. First wash buffer (need 150.0 mL): 50 mM HEPES– NaOH pH 7.5, 0.3 M NaCl, 1 mM EDTA–NaOH pH 7.5, 0.1% (v/v) NP-40. 16. Second wash buffer (need 25.0 mL): 50 mM HEPES– NaOH pH 7.5, 0.3 M NaCl, 1 mM EDTA–NaOH pH 7.5, 0.1% (v/v) NP-40, 10 mM β-ME.
- 25. Protein Antigen Expression in E. coli for Antibody Production 9 17. Avanti J-E centrifuge with JA-20 and JA-14 rotors (Beckman-Coulter, Fullerton, CA). 18. 250-mL Polycarbonate centrifuge bottles and 50-mL poly- carbonate centrifuge tubes. 19. Tabletop swinging bucket clinical centrifuge (Sorvall- Thermo Fisher Scientific, Waltham, MA). 2.5. Protease Removal of GST from Purified GST-Fusion Proteins 1. Purified GST-fusion protein (from Section 3.4.2). 2. Glutathione–SepharoseTM 4 Fast-Flow in TBS pH 7.4 at 25% (v/v). 3. Phosphate-buffered saline (PBS; 10× strength): Mix 80.0 g NaCl, 2.0 g KCl, 21.6 g Na2HPO4-7H2O, 2.0 g KH2PO4, and dH2O for a final volume of 1 L, sterile filter; use at 1× strength, sterile filter, and store at room temperature. 4. ProTEV Protease (Promega Corp., Madison, WI) (see Note 3). 5. Ni–SepharoseTM 6 Fast-Flow (GE Life-Sciences, Inc, Piscat- away, NJ); washed and equilibrated in PBS. 6. Thrombin (Novagen-EMD Biosciences, Madison, WI). 7. p-Aminobenzamidine–agarose (Sigma-Aldrich, St. Louis, MO). 3. Methods 3.1. SDS-PAGE 1. Attach precast gel to electrophoresis apparatus. Fill upper and lower reservoirs with 1× strength running buffer. 2. Heat samples at 65◦C for 15 min. Collect solution by cen- trifugation at 16,000×g for 5 min at room temperature. 3. Load 10 μL of each sample and protein molecular weight markers into individual wells. 4. Run gel according to manufacturer’s suggested cur- rent/voltage. 5. Disassemble gel apparatus and transfer gel carefully to Coomassie stain in a clean plastic/glass dish and cover. Incu- bate gel for 30 min (minimum) with gentle orbital shaking. 6. Decant stain and save for later use. Add both destain solution and several wadded kimwipes to the gel containing dish and cover. Incubate with gentle orbital shaking until sufficient contrast is observed between protein bands and gel back- ground. Frequently exchange used kimwipes for new ones to speed destaining.
- 26. Other documents randomly have different content
- 27. unless I be naked, Mistress whore!"[165] So she loosed her petticoat- trousers and, putting her hand under her clothes, tore them off her body; then twisted up a silken kerchief into cord-shape, girt it round her middle and became as she were a scald-head Ifritah or a spotted snake. With this she inclined towards the damsel and said, "Do thou as I have done." All this time, Sharrkan was gazing at the twain, and laughing at the beldam's loathly semblance. So the damsel leisurely rose and, taking a sash of Yamani stuff, passed it twice round her waist, then she tucked up her trousers and displayed two calves of alabaster carrying a mound of crystal, smooth and rounded, and a stomach which exhaled musk from its dimples, as it were a bed of Nu'uman's anemones; and breasts like double pomegranates. Then the old woman leant towards her, and the two laid hold either of each, while Sharrkan raised his head Heavenwards and prayed Allah that the belle might beat the beldam. Presently the young woman gat beneath the old woman; and, gripping her waist-cloth with the left and circling her neck with the right hand, hoisted her off the ground with both; whereupon the old woman strove to free herself and, in so doing fell on her back arsiversy, with her legs high in air and her hairy bush between them showed manifest in the moonshine; furthermore she let fly two great farts[166] one of which blew up the dust from the earth's face and the other steamed up to the gate of Heaven. Sharrkan laughed till he fell back upon the ground. Then he arose and, baring his brand looked right and left, but he saw no one save the old woman sprawling on her back, and said to himself, "He lied not who named thee Lady of Calamities! Verily thou knewest her prowess by her performance upon the others." So he drew near them to hear what should pass between them. Then the young lady went up to the old one and, throwing a wrapper of thin silk upon her nakedness, helped her to don her clothes and made excuses saying, "O my lady Zat al-Dawahi, I intended only to throw thee and not all this, but thou triedst to twist out of my hands; so laud to Allah for safety!" She returned her no answer, but rose in her shame and walked away till out of sight, leaving the handmaids prostrate and pinioned, with the fair damsel standing amongst them. Quoth Sharrkan to himself, "Every luck hath
- 28. its cause. Sleep did not fall upon me nor the war-horse bear me hither save for my good fortune; for doubtless this maid and what is with her shall become booty to me." So he made towards his steed and mounted and heeled[167] him on, when he sped as the shaft speeds from the bow, and in his hand he still hent his brand bare of sheath, which he brandished shouting the while his war-cry, "Allah is All-mighty[168] !" When the damsel saw him she sprang to her feet and, taking firm stand on the bank of the stream, whose breadth was six ells, the normal cubits, made one bound and landed clear on the farther side,[169] where she turned and cried out with a loud voice, "Who art thou, O thou fellow, that breakest in upon our privacy and pastime, and that too hanger in hand as if charging a host? Whence camest thou and whither art thou going? Speak sooth, for truth will stand thee in good stead, and lie not, for lies come of villein-breed. Doubtless thou hast wandered this night from thy way, that thou chancedst upon this place whence escape were the greatest of mercies; for thou art now in an open plain and, did we shout but a single shout, would come to our rescue four thousand knights.[170] So tell me what thou wantest; and if thou wouldst only have us set thee on the right road, we will do so." When Sharrkan heard her words he replied, "I am a stranger of the Moslems, who fared forth this night single-handed, seeking for spoil; nor could this moonlight show me a fairer booty than these ten maidens; so I shall seize them and rejoin my comrades with them." Quoth she, "I would have thee know that as for the booty thou hast not come at it; and, as for the handmaids, by Allah, they shall never be thy spoil. Have I not told thee that to lie is villein-vile?" Quoth he, "The wise man is he who taketh warning by others." Thereupon quoth she, "By the truth of the Messiah, did I not fear that thy death would be on my hands, I would shout a shout should fill the mead for thee with war steeds and with men of might, but I take pity upon the stranger. So, if thou seek booty, I require of thee that thou alight from thy steed and swear to me, by thy faith, that thou wilt not advance against me aught like arms in hand, and we will wrestle, I and thou. If thou throw me, set me on thy steed and take all of us to thy booty; but if I throw thee, thou shalt become under my
- 29. command. Swear this to me, for I fear thy treachery: indeed it hath become a common saw, Where Perfidy is innate there Trust is a weakly mate. Now an thou wilt swear I will return and draw near to thee and tackle thee." Answered Sharrkan (and indeed he lusted to seize her and said in his soul, "Truly she knoweth not that I am a champion of champions"); "Swear me by what oath thou wilt and by what thou deemest most binding, and I will not approach thee with aught till thou hast made thy preparation and sayest:—Draw near that I wrestle with thee. If thou throw me, I have money wherewithal to ransom myself; and if I throw thee, 'twill be booty and booty enough for me!" Rejoined the damsel, "I am content herewith!" and Sharrkan was astounded at her words and said, "And by the truth of the Apostle (whom Allah bless and keep!) I too am content on the other part!" Then said she, "Swear to me by Him who sprite in body dight and dealt laws to rule mankind aright, that thou wilt not offer me aught of violence save by way of wrestling; else mayst thou die without the pale of Al-Islam." Sharrkan replied, "By Allah! were a Kazi to swear me, even though he were a Kazi of the Kazis,[171] he would not impose upon me such an oath as this!" Then he sware to her by all she named and tied his steed to a tree; but he was drowned in the sea of thought, saying in himself, "Praise be to Him who fashioned her from dirty water!"[172] Then he girt himself and made ready for wrestling, and said to her, "Cross the stream to me;" but she replied, "It is not for me to come over to thee: if thou wilt, pass thou over here to me." "I cannot do that," quoth he, and quoth she, "O boy, I will come across to thee." So she tucked up her skirts and, leaping, landed on the other side of the stream by his side; whereupon he drew near to her and bent him forwards and clapped palms.[173] But he was confounded by her beauty and loveliness; for he saw a shape which the Hand of Power had tanned with the dye-leaves of the Jánn, which had been fostered by the Hand of Beneficence and fanned by the Zephyrs of fair fortune and whose birth a propitious ascendant had greeted. Then she called out to him, "O Moslem, come on and let us wrestle ere the break of morning," and tucked up her sleeves from a fore-arm like fresh curd, which illumined the whole place with its whiteness; and Sharrkan
- 30. was dazzled by it. Then he bent forwards and clapped his palms by way of challenge, she doing the like, and caught hold of her, and the two grappled and gripped and interlocked hands and arms. Presently he shifted his hands to her slender waist, when his finger tips sank into the soft folds of her middle, breeding languishment, and he fell a trembling like the Persian reed in the roaring gale. So she lifted him up and, throwing him to the ground, sat upon his breast with hips and hinder cheeks like mounds of sand, for his soul had lost mastery over his senses. Then she asked him, "O Moslem! the slaying of Nazarenes is lawful to you folk; what then hast thou to say about being slain thyself?"; and he answered, "O my lady, thy speech as regards slaying me is not other than unlawful; for our prophet Mohammed (whom Allah bless and preserve!) prohibited the slaying of women and children, old men and monks!" "As it was thus revealed to your Prophet," she replied, "it behoveth us to render the equivalent of his mercy; so rise. I give thee thy life, for generosity is never lost upon the generous." Then she got off his breast and he rose and stood shaking the dust from his head against the owners of the curved rib, even women; and she said to him, "Be not ashamed; but verily one who entereth the land of Roum in quest of booty, and cometh to assist Kings against Kings, how happeneth it that he hath not strength enough to defend himself from one made out of the curved rib?" "'Twas not for lack of strength in me," he answered; "nor didst thou throw me by thy force; it was thy loveliness overthrew me; so if thou wilt grant me another bout, it will be of thy courtesy." She laughed and said, "I grant thee thy request: but these handmaids have long been pinioned and their arms and sides are weary, and it were only right I should loose them, for haply this next wrestling bout will be long." Then she went to the slave-girls and, unbinding them, said to them in the tongue of Greece, "Get ye to some safe place, till I foil this Moslem's lust and longing for you." So they went away, whilst Sharrkan kept gazing at them and they kept turning to look at the two. Then each approached the adversary and he set his breast against hers, but when he felt waist touch waist, his strength failed him; and she, waxing ware of this, lifted him with her hands swiftlier than the blinding leven-flash, and threw him to
- 31. the ground. He fell on his back,[174] and then she said to him, "Rise: I give thee thy life a second time. I spared thee in the first count because of thy Prophet, for that he made unlawful the slaying of women; and I do so on the second count because of thy weakliness and the greenness of thine years and thy strangerhood; but I charge thee, if there be in the Moslem army sent by Omar bin al-Nu'uman to succour the King of Constantinople, a stronger than thou, send him hither and tell him of me: for in wrestling there are shifts and trips, catches and holds, such as the feint or falsing and the snap or first grip, the hug, the feet-catch, the thigh-bite,[175] the jostle and the leg-lock." "By Allah, O my lady," quoth Sharrkan (and indeed he was highly incensed against her), "had I been Master al-Safdí, Master Mohammed Kimál or Ibn al-Saddí,[176] as they were in their prime, I had kept no note of these shifts thou mentionest; for O my mistress, by Allah, thou hast not grassed me by thy strength, but by the blandishments of thy back-parts; for we men of Mesopotamia so love a full-formed thigh that nor sense was left me nor foresight. But now, an thou wish, thou shalt try a third fall with me while my wits are about me, and this last match is allowed me by the laws of the game which sayeth the best of three: moreover I have regained my presence of mind." When she heard his words she said to him, "Hast thou not had a belly-full of this wrestling, O vanquished one? However come on, an thou wilt; but know that this must be the last round." Then she bent forward and challenged him and Sharrkan did likewise, setting to it in real earnest and being right cautious about the throw: so the two strove awhile and the damsel found in him a strength such as she had not observed before and said to him, "O Moslem, thou art now on thy mettle." "Yes," he replied, "thou knowest that there remaineth to me but this one round, after which each of us will wend a different way." She laughed and he laughed too;[177] then she overreached at his thigh and caught firm hold of it unawares, which made him greet the ground and fall full on his back. She laughed at him and said, "Art thou an eater of bran? Thou art like a Badawi's bonnet which falleth off with every touch or else the Father of Winds[178] that droppeth before a puff of air. Fie upon thee, O thou poor thing!" adding, "Get thee back to the Moslem
- 32. army and send us other than thyself, for thou failest of thews; and proclaim for us, among the Arabs and Persians, the Turks and Daylamites,[179] whoso hath might in him, let him come to us." Then she made a spring and landed on the other side of the stream and said to Sharrkan, laughing, "Parting with thee is right grievous to me, O my lord; but get thee to thy mates before dawn, lest the Knights come upon thee and pick thee up on their lance-points. Thou hast no strength to defend thee against a woman, so how couldst thou hold thine own amongst men of might and Knights?" Sharrkan was confounded and called to her (as she turned from him making towards the convent), "O my lady, wilt thou go away and leave the miserable stranger, the broken hearted slave of love?" So she turned to him laughing and said, "What is thy want? I will grant thee thy prayer." "Have I set foot in thy country and tasted the sweetness of thy courtesy," replied he, "and shall I return without eating of thy victual and tasting thy hospitality; I who have become one of thy servitors!" "None baulk kindliness save the base," she rejoined, "honour us in Allah's name, on my head and eyes be it! Mount thy steed and ride along the brink of the stream over against me, for now thou art my guest." At this Sharrkan was glad and, hastening back to his horse, mounted and walked him abreast of her, and she kept faring on till they came to a drawbridge[180] built of beams of the white poplar, hung by pullies and steel-chains and made fast with hooks and padlocks. When Sharrkan looked, he saw awaiting her upon the bridge the same ten handmaids whom she had thrown in the wrestling-bouts; and, as she came up to them, she said to one in the Greek tongue, "Arise and take the reins of his horse and conduct him across into the convent." So she went up to Sharrkan and led him over, much puzzled and perturbed with what he saw, and saying to himself, "O would that the Wazir Dandan were here with me that his eyes might look upon these fairest of favours." Then he turned to the young lady and said to her, "O marvel of loveliness, now I have two claims upon thee; first the claim of good- fellowship, and secondly for that thou hast carried me to thy home and offered me thy hospitality. I am now under thy commandance and thy guidance; so do me one last favour by accompanying me to
- 33. the lands of Al-Islam; where thou shalt look upon many a lion- hearted warrior and thou shalt learn who I am." When she heard this she was angered and said to him, "By the truth of the Messiah, thou hast proved thyself with me a man of keen wit; but now I see what mischief there is in thy heart, and how thou canst permit thyself a speech which proveth thy traitorous intent. How should I do as thou sayest, when I wot that if I came to that King of yours, Omar bin al-Nu'uman, I should never get free from him? For truly he hath not the like of me or behind his city walls or within his palace- halls, Lord of Baghdad and of Khorasan though he be, who hath built for himself twelve pavilions, in number as the months of the year, and in each a concubine after the number of the days; and if I come to him he would not prove shy of me, for your folk believe I am lawful to have and to hold as is said in your writ:—Or those women whom your right hand shall possess as slaves.[181] So how canst thou speak thus to me? As for thy saying:—Thou shalt look upon the braves of the Moslems, by the truth of the Messiah, thou sayest that which is not true, for I saw your army when it reached our land, these two days ago; and I did not see that your ordinance was the ordinance of Kings, but I beheld only a rabble of tribesmen gathered together. And as to thy words:—Thou shalt know who I am, I did not do thee kindness because of thy dignity but out of pride in myself; and the like of thee should not talk thus to the like of me, even wert thou Sharrkan, Omar bin al-Nu'uman's son, the prowest name in these days!" "Knowest thou Sharrkan?" asked he; and she answered "Yes! and I know of his coming with an army numbering ten thousand horsemen; also that he was sent by his sire with this force to gain prevalence for the King of Constantinople." "O my lady," said Sharrkan, "I adjure thee by thy religion, tell me the cause of all this, that sooth may appear to me clear of untruth, and with whom the fault lies." "Now by the virtue of thy faith," she replied, "did I not fear lest the news of me be bruited abroad that I am of the daughters of Roum, I would adventure myself and sally forth single-handed against the ten thousand horsemen and slay their leader, the Wazir Dandan and vanquish their champion Sharrkan.[182] Nor would aught of shame accrue to me thereby, for I
- 34. have read books and studied the rules of good breeding in the language of the Arabs. But I have no need to vaunt my own prowess to thee, more by token as thou hast proved in thy proper person my skill and strength in wrestling; and thou hast learnt my superiority over other women. Nor, indeed, had Sharrkan himself been here this night and it were said to him:—Clear this stream, could he have done it; and I only long and lust that the Messiah would throw him into my hands in this very convent, that I might go forth to him in the habit of a man and drag him from his saddle-seat and make him my captive and lay him in bilboes."——And Shahrazad perceived the dawn of day and ceased to say her permitted say.
- 35. Now when it was the Forty-eighth Night, She said, It hath reached me, O auspicious King, that when the Nazarene damsel said to Sharrkan (and he listening impatiently
- 36. enow), "Verily if Sharrkan fell into my hands, I would go forth to him in the habit of a man and drag him from his saddle-seat and make him my captive and lay him in bilboes," pride and passion and knightly jealousy took possession of him and he desired to discover and declare himself and to lay on load; but her loveliness restrained him and he began repeating:— An faulty of one fault the Beauty prove, ✿ Her charms a thousand advocates shall move. So she went up and Sharrkan after her; and, when he saw the maiden's back and hinder cheeks that clashed against each other, like rollers in the rolling sea, he extemporised these couplets:— For her sins is a pleader that brow, ✿ And all hearts its fair pleading must trow: When I saw it I cried, "To-night ✿ The moon at its fullest doth show; Tho' Balkís' own Ifrit[183] try a bout, ✿ Spite his force she would deal him a throw. The two fared on till they reached a gate over which rose a marble archway. This she opened and ushered Sharrkan into a long vestibule, vaulted with ten connected arches, from each of which hung a crystal lamp glistening like a spark of fire. The handmaids met her at the further end bearing wax candles of goodly perfume, and wearing on their heads golden fillets crusted with all manner bezel-gems,[184] and went on before her (Sharrkan still following), till they reached the inner convent. There the Moslem saw couches and sofas ranged all around, one opposite the other and all overhung with curtains flowered in gold. The monastery floor was paved with every kind of vari-coloured marbles and mosaic-work, and in the midst stood a basin that held four-and-twenty jetting fountains of gold, whence the water ran like molten silver; whilst at the upper end stood a throne spread with silks fit only for Kings. Then said the damsel, "Ascend, O my lord, this throne." So he went up to it and sat down and she withdrew to remain absent for some time. Sharrkan asked of her from one of the servants who answered him, "She hath gone to her dormitory; but we will serve thee even as she ordered." So they set before him viands of rare varieties, and he ate
- 37. his sufficiency, when they brought him a basin of gold and an ewer of silver, and he washed his hands. Then his thoughts reverted to his army, knowing not what had befallen it in his absence and calling to mind also how he had forgotton his father's injunctions: so he was troubled about his case, repenting of what he had done till the dawn broke and the day appeared; when he lamented and sighed and became drowned in the sea of sadness and repeated:— I am not lost to prudence, but indeed ✿ Here I'm bewildered, what shall be my rede? Would any aid me in mine ails of love, ✿ By my own might and sleight would I be free'd: But ah! my heart is lost and passion-shent: ✿ To none save Allah can I trust my need! When he ended his verse behold, there came up to him a rare show and a fair, more than twenty maidens like crescents encompassing the young lady, who shone in their midst as the full moon among the constellations guarding and girding her. She was clad in brocades befitting Kings; her breasts were like twin pomegranates, a woven zone set with all kinds of jewels tightly clasped her waist which expanded below into jutting hips; and her hinder cheeks stood out as a mound of crystal[185] supporting a silvern shaft. When Sharrkan looked at her his wits went nigh to fly away from him with delight; and he forgot army and Wazir as he gazed on her fair head decked and dight with a net-work of pearls set off by divers sorts of gems. Handmaids on her right and handmaids on her left bore her train, as she paced with dainty graceful gait in all the pride of seemlihead. He sprang to his feet seeing such beauty and loveliness, and cried aloud, "Beware and beware of that zone rarely fair!" and broke out into these couplets:—
- 38. With heavy back-parts, high breasts delicate, ✿ And lissome form that sways with swimming gait, She deftly hides love-longing in her breast; ✿ But I may never hide its ban and bate: While hosts of followers her steps precede,[186] ✿ Like pearls now necklaced and now separate. She gazed upon him for a long time and considered him till she was assured of him, when she came up to him and said, "In very sooth the place is honoured and illumined by thee, O Sharrkan! How sped thy night, O hero, after we went away and left thee?"; adding, "Verily lying is a vile thing and a shameful, especially in great Kings! and thou art Crown-Prince Sharrkan, son and heir of King Omar bin al-Nu'uman; so henceforth make no secret of thy rank and condition, nor let me hear aught from thee but the truth; for leasing bequeatheth hate and despite. And as thou art pierced by the shaft of Fate, be resignation thine and abide content to wait." When he heard her words he saw that artifice availed him naught and he acknowledged the truth, saying, "I am Sharrkan, bin Omar bin al- Nu'uman, whom fortune hath afflicted, and cast into this place; so whatso thou willest, do it in my case!" She hung her head groundwards a long while, then turned to him and said, "Be of good cheer and let thine eyes be cool and clear;[187] for thou art the guest of my hospitality, and bread-and-salt hath made a tie between me and thee; wherefore thou art in my ward and under my safeguard. Have no fear for, by the truth of the Messiah, if all on earth sought to do thee hurt they should not come at thee, till life had left my body for thy sake: indeed thou art now under the charge of the Messiah and of me." Hereat she sat her down by his side and fell to playing with him, till his alarm subsided and he knew that had she desired to slay him, she would have done so during the past night. Presently she bespoke in the Grecian tongue one of her slave-girls, who went away and soon came back bringing a beaker and a tray of food; but Sharrkan abstained from eating and said to himself, "Haply she hath put somewhat in this meat." She knew what was in his thought; so she turned to him and said, "By the truth of the
- 39. Messiah, the case is not on such wise, nor is there aught in this meat of what thou suspectest! Had my mind been set on slaying thee, I had slain thee ere now." Then she walked up to the tray and ate of every dish a mouthful; whereupon Sharrkan came forward and ate too. She was pleased at this and both ate till they were satisfied. They washed their hands and after that she rose and ordered a handmaid to bring perfumes and herbs of sweet savour, wines of all colours and kinds and a wine-service with vessels of gold, silver and crystal. She filled a first goblet and drank it off before offering it to him, even as she had done with the food: then she downed a second and handed it to him. He drank and she said to him, "O Moslem, see how thou art here in all solace and delight of life!" And she ceased not to drink and ply him with drink, till he took leave of his wits,——And Shahrazad perceived the dawn of day, and ceased saying her permitted say. Now when it was the Forty-ninth Night, She said, It hath reached me, O auspicious King, that the damsel ceased not to drink and ply Sharrkan with drink till he took leave of his wits, for the wine and the intoxication of love he bore her. Presently she said to the slave-girl, "O Marjánah[188] ! bring us some instruments of music!" "To hear is to obey," said the handmaid and going out, returned in the twinkling of an eye with a Damascus lute, [189] a Persian harp, a Tartar pipe, and an Egyptian dulcimer. The young lady took the lute and, after tuning each several string, began in gentle undersong to sing, softer, than zephyr's wing and sweeter than Tasmin[190] -spring, with heart safe and secure from everything the couplets following:—
- 40. Allah assain those eyne! What streams of blood they shed! ✿ How many an arrowy glance those lids of thine have sped. I love all lovers who to lovers show them dure; ✿ Twere wrong to rue the love in wrong-head born and bred: Haply fall hapless eye for thee no sleeping kens! ✿ Heaven help the hapless heart by force of thee misled! Thou doomest me to death who art my king, and I ✿ Ransom with life the deemster who would doom me dead. Thereupon each and every of the maidens rose up and taking an instrument, played and recited couplets in the Roumi tongue; then their mistress sang also and seeing Sharrkan in ecstasies asked him, "O Moslem, dost thou understand what I say?"; and he answered, "Nay, my ecstasy cometh from the beauty of thy finger-tips." She laughed and continued, "If I sing to thee in Arabic what wouldst thou do?" "I should no longer," quoth he, "be master of my senses." Then she took an instrument and, changing the measure, began singing these verses:— The smack of parting's myrrh to me, ✿ How, then, bear patience' aloë? I'm girt by ills in trinity ✿ Severance, distance, cruelty! My freedom stole that fairest she, ✿ And parting irks me bitterly. When she ended her verse, she looked at Sharrkan and found him lost to existence, and he lay for a while stretched at full length and prone among the maidens.[191] Then he revived and, remembering the songs, again inclined to mirth and merriment; and the twain returned to their wine and wassail, and continued their playing and toying, their pastime and pleasure till day ceased illuminating and night drooped her wing. Then the damsel went off to her dormitory and when Sharrkan asked after her they answered, "She is gone to her sleeping-chamber," whereto he rejoined, "Under Allah's ward and His good guard!" As soon as it was morning, a handmaid came to him and said to him, "My mistress biddeth thee to her." So he rose and followed her and, as he drew near her lodging, the damsels welcomed him with smitten tabrets and songs of greeting, and led him through a great door of ivory studded with pearls and jewels.
- 41. Thence they passed with him into a tall and spacious hall, at the upper end of which was a wide daïs carpeted with all kinds of silks, and round it open lattices commanding a view of trees and streams. About the saloon were figures carved in human form, and fashioned on such wise that the air passed through them and set in motion musical instruments within, so that the beholder would fancy they spoke.[192] Here sat the young lady, looking at the figures; but when she saw Sharrkan, she sprang to her feet and, taking him by the hand, made him sit down by her side, and asked him how he had passed the night. He blessed her and the two sat talking awhile till she asked him, "Knowest thou aught touching lovers and slaves of love?"; and he answered "Yes! I wot somewhat in verse on that matter." "Let me hear it," quoth she, so he began quoting:— Pleasure and health, good cheer, good appetite ✿ To Azzah, freest with our name and fame! By Allah! would I near her off she flies ✿ At tangent, granting less the more I claim: I dote on Azzah, but when clear I off ✿ My rivals, clears me too that dearest dame; Like wandering wight that chose for shade a cloud ✿ Which, ere siesta done, thin air became. When she heard this she said, "Verily Al-Kuthayyir[193] was conspicuous for sweet speech and chaste, and he was superlative in his praise of Azzah when he sang (and she began to recite):— "Did Azzah deal behest to Sun o' noon, ✿ The judge had 'judged her beauty's bestest boon; And girls who come to me and carp at her, ✿ God make their rosy cheeks her sandal-shoon!" "And indeed," quoth she, "'twas said that Azzah boasted exceeding beauty and loveliness." Then she asked Sharrkan saying, "O Prince, dost thou know aught of Jamíl's[194] verses to Buthaynah? if so repeat to us somewhat of them;" and he answered, "Yes, I know them better than any;" whereupon he began repeating these couplets:—
- 42. "Jamíl, in Holy war go fight!" to me they say: ✿ What war save fight for fair ones would I e'er essay? To me their every word and work are mere delight, ✿ And martyrs clepe I all they slay in fight and fray: An ask I, "O Buthaynah! what's this love, I pray, ✿ Which eats my heart? quoth she "'Twill stay for ever and aye!" And when I cry, "Of wits return some small display ✿ For daily use," quoth she, "Far, far 'tis fled away!" Thou seekst my death; naught else thy will can satisfy ✿ While I no goal espy save thee and thee alway. "Thou hast spoken right well," said she, "O King's son, and Jamíl also spoke excellently well. But what would Buthaynah have done with him that he saith in his hemistich:— Thou seekst my death; naught else thy will can satisfy?" "O my lady," quoth Sharrkan, "she willed to do him what thou willest to do with me, and even that will not satisfy thee." She laughed at his opportune reply and they ceased not carousing till Day put out her light and Night came in darkness dight. Then she rose and went to her dormitory and slept, while Sharrkan slept in his place till morning dawned. As soon as he awoke, the handmaids came to him with tabrets and other instruments of mirth and merriment, as wont; and, kissing the ground between his hands, said to him, "Bismillah! —in Allah's name—be so kind as to come[195] : our mistress biddeth thee to her presence!" So he rose and accompanied the slave-girls who surrounded him, playing on tabrets and other instruments of music, till they passed from that saloon into another and a yet more spacious hall, decorated with pictured likenesses and figures of birds and beasts, passing all description. Sharrkan marvelled at the art and artifice of the place and began reciting:—
- 43. He pluckt fruits of her necklace in rivalry, ✿ And her breast-pearls that bedded in gold-mine lie. Pure water on silvern bars is her brow, ✿ And her cheeks show roses with rubies vie: Meseems in her eyne that the violet's hue ✿ Lies purpling set in the Ithmid's[196] dye. When the lady saw Sharrkan, she stood up to him in honour and, taking his hand, seated him by her side and asked, "O son of King Omar bin al-Nu'uman, hast thou any cunning in the game of chess?" "Yes," he answered, "but do not thou with me as said the poet:— I speak and longing love upties me and unties me; ✿ Till with her honey-dew of inner lip she plies me: I brought the chess-board and my liefest lover plays me ✿ With white and black, [197] but black-cum-white ne'er satisfies me: 'Twas as if King for Castle I were fain to place me ✿ Till wilful loss of game atwixt two queens surprise me: And if I seek to read intent in eyes that eye me ✿ Oh man! that glance askance with hint of wish defies me." Then she brought the chess-board and played with him; but Sharrkan, instead of looking at her moves, kept gazing at her fair mouth, and putting knight in place of elephant and elephant[198] in stead of knight. She laughed and said to him, "If thy play be after this fashion, thou knowest naught of the game." "This is only our first," replied he, "judge not by this bout." When she beat him he replaced the pieces in position and played again with her; but she beat him a second time, a third, a fourth and a fifth. So she turned to him and said, "Thou art beaten in everything;" and he replied, "O my lady, how should one playing with the like of thee avoid being beaten?" Then she bade bring food, and they ate and washed their hands; after which the wine was set before them and they drank. Presently, she took the dulcimer, for her hand was cunning in smiting it, and she began repeating to an accompaniment these couplets:—
- 44. Twixt the close-tied and open-wide no medium Fortune knoweth; ✿ Now ebb and flow then flow and ebb this wise her likeness showeth: Then drink her wine the syne she's thine and smiling thou dost find her; ✿ Anon she'll fall and fare away when all thy good forth goeth. They ceased not to carouse till nightfall and this day was pleasanter even than the first. When darkness set in, the lady betook her to her dormitory, leaving him alone with the handmaids; so he threw himself on the ground and slept till dawn, when the damsels came to him with tambourines and other instruments according to custom. Seeing them he roused him hastily and sat up; and they carried him to their mistress, who came to meet him and, taking him by the hand, seated him by her side. Then she asked him how he had passed his night, whereat he prayed that her life be prolonged; and she took the lute and sang to it these verses which she improvised: — Ne'er incline thee to part ✿ Which embitters the heart; E'en the sun when he sets ✿ Shall in pallor depart. While they were solacing themselves after this fashion, behold, there arose a great and sudden clamour, and a confused crowd of knights and men rushed in, holding drawn swords that glittered and gleamed in their hands, and cried aloud in the Grecian tongue, "Thou hast fallen into our hands, O Sharrkan, so make thee sure of death!" When he heard this, he said to himself, "By Allah, she hath entrapped me and held me in play, till her men should come. These are the Knights with whom she threatened me; but 'tis I who have thrown myself into this strait." Then he turned towards the young lady to reproach her, but saw that she had changed colour and her face was pale; and she sprang to her feet and asked the crowd, "Who are ye?" "O most gracious Princess and peerless union-pearl," answered the leading Knight, "dost thou weet who is yon man by thy side?" "Not I," she replied, "who may he be?" Quoth the Patrician, "This is of towns the highwayman! This is he who rideth in the horseman's van! This is Sharrkan, son of King Omar bin al- Nu'uman! This is he that forceth fortalice and penetrateth every
- 45. impregnable place! The news of him reached King Hardub, thy father, by report of the ancient dame Zat al-Dawahi; and thy sire, our sovereign, hath made sure that thou hast rendered good service to the army of the Greeks by taking captive this ominous lion." When she heard this, she looked at the Knight and asked him, "What be thy name?" and he answered, "I am Masurah, son of thy slave Mausúrah bin Káshardah, Knight of Knights." "And how?" quoth she, "durst thou enter my presence without leave?" Quoth he, "O my lady, when I came to the gate, none forbade me, neither chamberlain nor porter, but all the doorkeepers rose and forewent us as of wont; although, when others come, they leave them standing at the gate while they ask permission to admit them. But this is not a time for long talking, when the King is expecting our return with this Prince, the scorpion-sting[199] of the Islamitic host, that he may kill him and drive back his men whither they came, without the bane of battling with them." "These words be ill words," rejoined the Princess, "and Dame Zat al-Dawahi lied, avouching an idle thing and a vain, whereof she weeteth not the truth; for by the virtue of the Messiah, this man who is with me is not Sharrkan, nor is he a captive, but a stranger who came to us seeking our hospitality, and I made him my guest. So even were we assured that this be Sharrkan and were it proved to us that it is he beyond a doubt, I say it would ill befit mine honour that I should deliver into your hands one who hath entered under my protection. So make me not a traitor to my guest and a disgrace among men; but return to the King, my father, and kiss the ground before him, and inform him that the case is contrariwise to the report of the Lady Zat al-Dawahi." "O Abrizah," replied Masurah, the Knight, "I cannot return to the King's majesty without his debtor and enemy." Quoth she (and indeed she had waxed very wroth), "Out on thee! Return to him with my answer, and no blame shall befal thee!" Quoth Masurah, "I will not return without him." Thereupon her colour changed and she exclaimed, "Exceed not in talk and vain words; for verily this man had not come in to us, were he not assured that he could of himself and single- handed make head against an hundred riders; and if I said to him:— Thou art Sharrkan, son of King Omar bin al-Nu'uman, he would
- 46. answer, Yes. But 'tis not of your competence to let or hinder him; for if you so do, he will not turn back from you till he hath slain all that are in this place. Behold, here he is by my side, and I will bring him before you sword and targe in hand." "Albeit I were safe from thy wrath," answered Masurah the Knight, "I am not safe from that of thy father, and when I see him, I shall sign to the Knights to take him captive, and we will carry him to the King bound and in abject sort." When she heard this, she said, "The matter shall not pass thus, for 'twould be blazoning mere folly. This man is but one and ye are an hundred Knights: so if you would attack him come out against him, one after one, that it may appear to the King which is the valiant amongst you."——And Shahrazad perceived the dawn of day and ceased to say her permitted say. Now when it was the Fiftieth Night, She said, It hath reached me, O auspicious King, that Princess Abrizah said to the Knight, "This man is but one, and ye are an hundred: so if ye would attack him, come out against him, one after one, that it may appear to the King which is the valiant." Quoth Masurah, the Knight, "By the truth of the Messiah, thou sayest sooth, and none but I shall sally out against him first." Quoth she, "Wait till I go to him and acquaint him with the case and hear what answer he will make. If he consent, 'tis well; but if he refuse, ye shall on no wise come to him, for I and my handmaids and whosoever is in the convent will be his ransom." So she went to Sharrkan and told him the news, whereat he smiled and knew that she had not informed any of the Emirs; but that tidings of him had been bruited and blazed abroad, till the report reached the King, against her wish and intent. So he again began reproaching himself and said, "How came I to adventure and play with my life by coming to the country of the Greeks?" But hearing the young lady's proposal he said to her, "Indeed their onset, one after one, would be overburdensome to them. Will they not come out against me, ten by
- 47. ten?" "That would be villeiny," said she; "Let one have at one." When he heard this, he sprang to his feet and made for them with his sword and battle-gear; and Masurah, the Knight, also sprang up and bore down upon him. Sharrkan met him like a lion and delivered a shoulder cut[200] which clove him to the middle, and the blade came out gleaming and glittering from his back and bowels. When the lady beheld that swashing blow, Sharrkan's might was magnified in her sight and she knew that when she overthrew him in the wrestle it was not by her strength but by her beauty and loveliness. So she turned to the Knights and said, "Take wreak for your chief!" Thereupon out came the slain man's brother, a fierce and furious Knight, and rushed upon Sharrkan, who delayed not, but smote him also with the shoulder-cut and the sword came out glittering from his vitals. Then cried the Princess, "O ye servants of the Messiah, avenge your comrade!" So they ceased not charging down upon him, one after one; and Sharrkan also ceased not playing upon them with the blade, till he had slain fifty knights, the lady looking on the while. And Allah cast a panic into the hearts of the survivors, so that they held back and dared not meet him in the duello, but fell upon him in a body; and he laid on load with heart firmer than a rock, and smote them and trod them down like straw under the threshing-sled, [201] till he had driven sense and soul out of them. Then the Princess called aloud to her damsels, saying, "Who is left in the convent?"; and they replied, "None but the gate-keepers;" whereupon she went up to Sharrkan and took him to her bosom, he doing the same, and they returned to the palace, after he had made an end of the melée. Now there remained a few of the Knights hiding from him in the cells of the monastery, and when the Princess saw this she rose from Sharrkan's side and left him for a while, but presently came back clad in closely-meshed coat of ring-mail and holding in her hand a fine Indian scymitar. And she said, "Now by the truth of the Messiah, I will not be a niggard of myself for my guest; nor will I abandon him though for this I abide a reproach and a byword in the land of the Greeks." Then she took reckoning of the dead and found that he had slain fourscore of the Knights, and other twenty had taken to flight.[202] When she saw what work he had made with them she said
- 48. to him, "Allah bless thee, O Sharrkan! The Cavaliers may well glory in the like of thee." Then he rose and wiping his blade clean of the blood of the slain began reciting these couplets:— How oft in the mellay I've cleft the array, ✿ And given their bravest to lions a prey: Ask of me and of them when I proved me prow ✿ O'er creation, on days of the foray and fray: When I left in the onslaught their lions to lie ✿ On the sands of the lowlands[203] in fieriest day. When he ended his verse, the Princess came up to him with smiles and kissed his hand; then she doffed her hauberk and he said to her, "O lady mine, wherefore didst thou don that coat of mail and bare thy brand?" "To guard thee against these caitiffs,"[204] she replied. Then she summoned the gate-keepers and asked them, "How came ye to admit the King's Knights into my dwelling without leave of me?"; and they answered, "O Princess, it is not our custom to ask leave of thee for the King's messengers, and especially for the chief of his Knights." Quoth she, "I think ye were minded only to disgrace me and murder my guest;" and bade Sharrkan smite their necks. He did so and she cried to the rest of her servants, "Of a truth, they deserved even more than that!" Then turning to Sharrkan, she said to him, "Now that there hath become manifest to thee what was concealed, thou shalt be made acquainted with my history. Know, then, that I am the daughter of King Hardub of Roum; my name is Abrizah and the ancient dame, yclept Zat al-Dawahi, is my grandmother by the sword side. She it certainly is who told my father of thee, and as surely she will compass a sleight to slay me, more by token as thou hast slain my father's chivalry and it is noised abroad that I have separated myself from the Nazarenes and have become no better than I should be with the Moslems. Wherefore it were wiser that I leave this dwelling while Zat al-Dawahi is on my track; but I require of thee the like kindness and courtesy I have shown thee, for enmity will presently befal between me and my father on thine account. So do not thou neglect to do aught that I shall say to thee, remembering all this betided me not save by
- 49. reason of thee." Hearing her words, Sharrkan joyed greatly; his breast broadened and his wits flew from him for delight, and he said, "By Allah, none shall come at thee, while life is in my bosom! But hast thou patience to bear parting from thy parents and thy people?" "Even so," she answered; and Sharrkan swore to her and the two plighted their troth. Then said she, "Now is my heart at ease; but there remaineth one other condition for thee." "What is it?" asked he and she answered, "It is that thou return with thy host to thine own country." Quoth he, "O lady mine, my father, King Omar bin al-Nu'uman, sent me to wage war upon thy sire, on account of the treasure he plundered from the King of Constantinople, and amongst the rest three great jewels, noted givers of good fortune." Quoth she, "Cheer thy heart and clear thine eyes: I will tell thee the whole of the tale and the cause of our feud with the King of Constantinople. Know that we have a yearly festival, hight the Convent-Feast, whereat Kings from all quarters and the noblest women are wont to congregate; thither also come merchants and traders with their wives and families, and the visitors abide there seven days. I was wont to be one of them; but, when there befel enmity between us, my father forbade me to be present at the festival for the space of seven years. One year, it chanced that amongst the daughters of the great who resorted to the patron, as was their custom, came a daughter of the King of Constantinople, a beautiful girl called Sophia. They tarried at the monastery six days and on the seventh the folk went their ways;[205] but Sophia said, I will not return to Constantinople save by water. So they equipped for her a ship in which she embarked with her suite; and making sail they put out to sea; but as they were voyaging behold, a contrary wind caught them and drove the vessel from her course till, as Fate and Fortune would have it, she fell in with a Nazarene craft from the Camphor Island[206] carrying a crew of five hundred armed Franks, who had been cruising about a long time. When they sighted the sails of the ship, wherein Sophia and her women were, they gave chase in all haste and in less than an hour they came up with her, when they laid the grappling-irons aboard her and captured her. Then taking her in tow they made all sail for their own island and
- 50. were but a little distant from it when the wind veered round and, splitting their sails, drove them on to a shoal which lies off our coast. Thereupon we sallied forth and, looking on them as spoil driven to us by Fate,[207] boarded and took them; and, slaying the men, made prize of the wreck, wherein we found the treasures and rarities in question and forty maidens, amongst whom was the King's daughter, Sophia. After the capture we carried the Princess and her women to my father, not knowing her to be a daughter of King Afridun of Constantinople; and he chose out for himself ten including her; and divided the rest among his dependants. Presently he set apart five damsels, amongst whom was the King's daughter, and sent them to thy father, King Omar bin al-Nu'uman, together with other gifts, such as broadcloth[208] and woollen stuffs and Grecian silks. Thy father accepted them and chose out from amongst the five girls Sophia, daughter of King Afridun; nor did we hear more of her till the beginning of this year, when her father wrote to my father in words unfitting for me to repeat, rebuking him with menaces and saying to him:—Two years ago, you plundered a ship of ours which had been seized by a band of Frankish pirates in which was my daughter Sophia, attended by her maidens numbering some threescore. Yet ye informed me not thereof by messenger or otherwise; nor could I make the matter public, lest reproach befal me amongst the Kings, by reason of my daughter's honour. So I concealed my case till this year, when I wrote to certain Frankish corsairs and sought news of my daughter from the Kings of the Isles. They replied:—By Allah we carried her not forth of thy realm; but we have heard that King Hardub rescued her from certain pirates. And they told me the whole tale. Then he added in the writing which he writ to my father:— Except you wish to be at feud with me and design to disgrace me and dishonour my daughter, you will, the instant my letter reacheth you, send my daughter back to me. But if you slight my letter and disobey my commandment, I will assuredly make you full return for your foul dealing and the baseness of your practices.[209] When my father read this letter and understood the contents,[210] it vexed him and he regretted not having known that Sophia, King Afridun's daughter, was among the captured damsels, that he might have sent
- 51. her back to her sire; and he was perplexed about the case because, after so long a time, he could not send to King Omar bin al-Nu'uman and demand her back from him, especially as he had lately heard that Heaven had granted him boon of babe by this Sophia. So when we pondered that truth, we knew that this letter was none other than a grievous calamity; and my father found nothing for it but to write an answer to King Afridun, making his excuses and swearing to him by strong oaths that he knew not his daughter to be among the bevy of damsels in the ship and setting forth how he had sent her to King Omar bin al-Nu'uman, who had gotten the blessing of issue by her. When my father's reply reached King Afridun he rose up and sat down,[211] and roared and foamed at the mouth crying:—What! shall he take captive my daughter and even her with slave-girls and pass her on from hand to hand sending her for a gift to Kings, and they lie with her without marriage-contract? By the Messiah and the true Faith, said he, I will not desist till I have taken my blood-vengeance for this and have wiped out my shame; and indeed I will do a deed which the chroniclers shall chronicle after me! So he bided his time till he devised a device and laid notable toils and snares, when he sent an embassy to thy father, King Omar, to tell him that which thou hast heard: accordingly thy father equipped thee and an army with thee and sent thee to King Afridun, whose object is to seize thee and thine army to boot. As for the three jewels whereof he told thy father when asking his aid, there was not one soothfast word in that matter, for they were with Sophia, his daughter; and my father took them from her, when he got possession of her and of her maidens, and gave them to me in free gift, and they are now with me. So go thou to thy host and turn them back ere they be led deep into, and shut in by, the land of the Franks and the country of the Greeks; for as soon as you have come far enough into their interior, they will stop the roads upon you and there will be no escape for you till the Day of retribution and retaliation. I know that thy troops are still halting where thou leftest them, because thou didst order a three days' rest; withal they have missed thee all this time and they wot not what to do." When Sharrkan heard her words, he was absent awhile in thought; then he kissed Princess Abrizah's hand and said,
- 52. "Praise be to Allah who hath bestowed thee on me and appointed thee to be the cause of my salvation and the salvation of whoso is with me! But 'tis grievous to me to part from thee and I know not what will become of thee after my departure." "Go now to thine army," she replied, "and turn them back, while ye are yet near your own country. If the envoys be still with them, lay hands on them and keep them, that the case may be made manifest to you; and, after three days, I will be with you all and we will enter Baghdad together." As he turned to depart she said, "Forget not the compact which is between me and thee;" then she rose to bid[212] him farewell and embrace him and quench the fire of desire, so she took leave of him and, throwing her arms round his neck, wept with exceeding weeping, and repeated these verses:— I bade adieu, my right hand wiped my tears away, ✿ The while my left hand held her in a close embrace: "Fearest thou naught," quoth she, "of shame?" I answered "Nay, ✿ The lover's parting day is lover's worst disgrace." Then Sharrkan left her and walked down from the convent. They brought his steed, so he mounted and rode down-stream to the drawbridge which he crossed and presently threaded the woodland paths and passed into the open meadow. As soon as he was clear of the trees he was aware of horsemen which made him stand on the alert, and he bared his brand and rode cautiously; but as they drew near and exchanged curious looks he recognized them and behold, it was the Wazir Dandan and two of his Emirs. When they saw him and knew him, they dismounted and saluting him, asked the reason of his absence; whereupon he told them all that had passed between him and Princess Abrizah from first to last. The Wazir returned thanks to Almighty Allah for his safety and said,[213] "Let us at once leave these lands; for the envoys who came with us are gone to inform the King of our approach, and haply he will hasten to fall on us and take us prisoners." So Sharrkan cried to his men to saddle and mount, which they did and, setting out at once, they stinted not faring till they reached the sole of the valley wherein the host lay.
- 53. The Ambassadors meanwhile had reported Sharrkan's approach to their King, who forthright equipped a host to lay hold of him and those with him. But Sharrkan, escorted by the Wazir Dandan and the two Emirs, had no sooner sighted the army, than he raised the cry "March! March!" They took horse on the instant and fared through the first day and second and third day, nor did they cease faring for five days; at the end of which time they alighted in a well-wooded valley, where they rested awhile. Then they again set out and stayed not riding for five and twenty days which placed them on the frontiers of their own country. Here, deeming themselves safe, they halted to rest; and the country people came out to them with guest- gifts for the men and provender and forage for the beasts. They tarried there two days after which, as all would be making for their homes, Sharrkan put the Wazir Dandan in command, bidding him lead the host back to Baghdad. But he himself remained behind with an hundred riders, till the rest of the army had made one day's march: then he called "To horse!" and mounted with his hundred men. They rode on two parasangs'[214] space till they arrived at a gorge between two mountains and lo! there arose before them a dark cloud of sand and dust. So they checked their steeds awhile till the dust opened and lifted, discovering beneath it an hundred cavaliers, lion-faced and in mail-coats cased. As soon as they drew within earshot of Sharrkan and his meiny they cried out to them, saying, "By the virtue of John and Mary, we have won to our wish! We have been following you by forced marches, night and day, till we forewent you to this place. So dismount and lay down your arms and yield yourselves, that we may grant you your lives." When Sharrkan heard this, his eyes stood out from his head and his cheeks flushed red and he said "How is it, O Nazarene dogs, ye dare enter our country and overmarch our land? And doth not this suffice you, but ye must adventure your selves and address us in such unseemly speech? Do you think to escape out of our hands and return to your country?" Then he shouted to his hundred horsemen, "Up and at these hounds, for they even you in number!" So saying, he bared his sabre and bore down on them, he and his, but the Franks met them with hearts firmer than rocks, and wight clashed against wight, and
- 54. knight dashed upon knight, and hot waxed the fight, and sore was the affright, and nor parley nor cries of quarter helped their plight; and they stinted not to charge and to smite, right hand meeting right, nor to hack and hew with blades bright-white, till day turned to night and gloom oppressed the sight. Then they drew apart and Sharrkan mustered his men and found none wounded save four only, who showed hurts but not death-hurts. Said he to them, "By Allah, my life long have I waded in the clashing sea of fight and I have met many a gallant sprite, but none so unfrightened of the sword that smites and the shock of men that affrights like these valiant Knights!" "Know, O King," said they, "that there is among them a Frankish cavalier who is their leader and, indeed, he is a man of valour and fatal is his spear-thrust: but, by Allah, he spares us great and small; for whoso falls into his hands he lets him go and forbears to slay him. By Allah, had he willed he had killed us all." Sharrkan was astounded when he heard what the Knight had done and such high report of him, so he said, "When the morn shall morrow, we will draw out and defy them, for we are an hundred to their hundred; and we will seek aid against them from the Lord of the Heavens." So they rested that night in such intent; whilst the Franks gathered round their Captain and said, "Verily this day we did not win our will of these;" and he replied, "At early dawn when the morrow shall morn, we will draw out and challenge them, one after one." They also rested in that mind, and both camps kept guard until Almighty Allah sent the light of day-dawn. Thereupon King Sharrkan and his hundred riders took horse and rode forth to the plain, where they found the Franks ranged in line of battle; and Sharrkan said to his followers, "Our foes have determined like ourselves to do their devoir; so up and at them and lay on load." Then came forth an Herald of the Franks and cried out, saying, "Let there be no general engagement betwixt us this day, save by the duello, a champion of yours against a champion of ours." Whereupon one of Sharrkan's riders dashed out from the ranks and drave between the two lines crying, "Ho! who is for smiting? Let no dastard engage me this day nor nidering!" Hardly had he made an end of his vaunt, when there sallied forth to him a Frankish cavalier,
- 55. armed cap-à-pie and clad in a surcoat of gold stuff, riding on a grey- white steed,[215] and he had no hair on his cheeks. He urged his charger on to the midst of the battle-plain and the two fell to derring-do of cut and thrust, but it was not long before the Frank foined the Moslem with the lance-point; and, toppling him from his steed, took him prisoner and led him off crestfallen. His folk rejoiced in their comrade and, forbidding him to go out again to the field, sent forth another, to whom sallied out another Moslem, brother to the captive, and offered him battle. The two fell to, either against other, and fought for a little while, till the Frank bore down upon the Moslem and, falsing him with a feint, tumbled him by a thrust of the lance-heel from his destrier and took him prisoner. After this fashion the Moslems ceased not dashing forwards, one after one, and the Franks to unhorse them and take them captive, till day departed and the night with darkness upstarted. Now they had captured of the Moslems twenty cavaliers, and when Sharrkan saw this, it was grievous to him and he mustered his men and said to them, "What is this thing that hath befallen us? To-morrow, I myself will go forth to the field and offer singular combat to their chief and learn what is the cause of his entering our land and warn him against doing battle with our band. If he persist, we will punish him with death, and if he prove peaceable we will make peace with him." They nighted on this wise till Allah Almighty caused the morn to dawn, when mounted the twain and drew up for battle fain; and Sharrkan was going forth to the plain, but behold, more than one half of the Franks dismounted and remained on foot before one of them who was mounted, till they reached the midst of the battle-plain. Sharrkan looked at that horseman and lo! he was their chief. He was clad in a surcoat of blue satin and a close-ringed mail shirt; his face was as the moon when it rises and no hair was upon his cheeks. He hent in hand an Indian scymitar and he rode a sable steed with a white blaze on brow, like a dirham; and he smote the horse with heel till he stood almost in the midst of the field when, signing to the Moslems, he cried out in fluent Arab speech, "Ho, Sharrkan! Ho, son of Omar bin al-Nu'uman! Ho, thou who forcest fortalice and overthrowest cities and countries! up and out to battle-bout, and blade single-handed wield with one
- 56. who halves with thee the field! Thou art Prince of thy people and I am Prince of mine; and whoso overcometh his adversary, him let the other's men obey and come under his sway." Hardly had he ended his speech, when out came Sharrkan with a heart full of fury, and urging his steed into the midst of the field, closed like a raging lion with the Frank who encountered him with wariness and steadfastness and met him with the meeting of warriors. Then they fell to foining and hewing, and they stinted not of onset and offset, and give and take, as they were two mountains clashing together or two seas together dashing; nor did they cease fighting until day darkened and night starkened. Then they drew apart and each returned to his own party; but as soon as Sharrkan foregathered with his comrades, he said, "Never looked I on the like of this cavalier: he hath one quality I have not yet seen in any and this it is that, when his foemen uncovereth a place for the death-blow, he reverseth his weapon and smiteth with the lance-heel! In very deed I know not what will be the issue 'twixt him and me; but 'tis my wish that we had in our host his like and the like of his men." Then he went to his rest for the night and, when morning dawned, the Frank came forth and rode down to the mid-field, where Sharrkan met him; and they fell to fighting and to wheeling, left and right; and necks were stretched out to see the sight, nor did they stint from strife and sword-play and lunge of lance with main and might, till the day turned to night and darkness overwhelmed the light. Then the twain drew asunder and returned each to his own camp, where both related to their comrades what had befallen them in the duello; and at last the Frank said to his men, "To-morrow shall decide the matter!" So they both passed that night restfully till dawn; and, as soon as it was day, they mounted and each bore down on other and ceased not to fight till half the day was done. Then the Frank bethought him of a ruse; first urging his steed with heel and then checking him with the rein, so that he stumbled and fell with his rider; thereupon Sharrkan threw himself on the foe, and would have smitten him with the sword fearing lest the strife be prolonged, when the Frank cried out to him, "O Sharrkan, champions are not wont to do thus! This is the act of a man accustomed to be beaten
- 57. by a woman."[216] When Sharrkan heard this, he raised his eyes to the Frank's face and gazing steadfastly at him, recognized in him Princess Abrizah with whom that pleasant adventure had befallen him in the convent; whereupon he cast brand from hand and, kissing the earth before her, asked her, "What moved thee to a deed like this?"; and she answered, "I desired to prove thy prowess afield and test thy doughtiness in tilting and jousting. These that are with me are my handmaids, and they are all clean maids; yet they have vanquished thy horsemen in fair press and stress of plain; and had not my steed stumbled with me, thou shouldst have seen my might and prowess in combat." Sharrkan smiled at her speech and said, "Praise be to Allah for safety and for my reunion with thee, O Queen of the age." Then she cried out to her damsels to loose the twenty captives of Sharrkan's troop and dismount. They did as she bade and came and kissed the earth before her and Sharrkan who said to them, "It is the like of you that Kings keep in store for the need- hour." Then he signed to his comrades to salute the Princess; so all alighted and kissed the earth before her, for they knew the story. After this, the whole two hundred took horse, and fared on night and day for six days' space, till they drew near to Baghdad, when they halted and Sharrkan bade Abrizah and her handmaids doff the Frankish garb that was on them——And Shahrazad perceived the dawn of day and ceased saying her permitted say. Now when it was the Fifty-first Night, She said, It hath reached me, O auspicious King, that Sharrkan bade Princess Abrizah and her damsels doff the garb that was on them and don the garments of daughters of Greece; and thus did they. Then he despatched a company of his companions to Baghdad to acquaint his father Omar bin al-Nu'uman, with his arrival and report that he was accompanied by Princess Abrizah, daughter of King Hardub, Lord of Græcia-land. They halted forthright in the place they had reached, and Sharrkan also halted and all nighted there; and
- 58. when Almighty Allah made morning dawn, Sharrkan and his company and Abrizah and her company took horse and fared on towards the city; when lo! on the way they met the Wazir Dandan, who had come out amongst a thousand horse, to honour Abrizah and Sharrkan, by especial commandment of King Omar Son of Al- Nu'uman. When the two drew near, they turned towards them and kissed ground before them; then they mounted again and escorted them into the city and went up with them to the palace. Sharrkan walked in to his father, who rose and embraced him and questioned him of his case. So he told him all that Abrizah had told him, and what had passed between them and said, "She hath parted from her sire and departed from her reign and hath chosen to take part with us and to make her abode with us; and indeed (he said to his father) the King of Constantinople hath plotted to do us a mischief, because of his daughter Sophia, for that the King of Greece had made known to him her story and the cause of her being given to thee; and he (the Grecian King) not knowing her to be daughter of King Afridun, Lord of Constantinople; and, had he known that, he would not have bestowed her upon thee, but he would have restored her to her parent. And of a verity (he continued) we were saved from these perils only by the Lady Abrizah, and never saw we a more valiant than she." And he went on to tell his father all that had passed from first to last of the wrestling and the single-fighting. When King Omar heard the story of Sharrkan, Abrizah was exalted in his eyes, and he longed to see her and question her. Thereupon Sharrkan went out to her and said, "The King calleth for thee;" she replied, "I hear and I obey;" and he took her and brought her in to his father, who was seated on his throne and who, having dismissed his high officers, was attended only by his eunuchs. The Princess entered and kissing the ground between his hands, saluted him in choice terms. He was amazed at her eloquent speech and thanked her for her dealing with his son Sharrkan and bade her be seated. So she sat down and unveiled her face;[217] and, when the King saw her beauty, his reason fled his head and he made her draw near and showed her favour, appointing her an especial palace for herself and her damsels, and assigning them solde and allowances. Then began
- 59. he to ask her of the three jewels aforesaid, and she answered, "Here be they with me, O King of the age!" So saying, she rose and going to her lodging, unpacked her baggage and from it brought out a box and from the box a casket of gold. She opened the casket and taking out those three jewels, kissed them and gave them to the King. Then she went away bearing his heart with her. After her going the King sent for his son Sharrkan and gave him one jewel of the three, and when he enquired of the other two replied, "O my son! I mean to give one to thy brother Zau al-Makan, and the other to thy sister Nuzhat al-Zaman." But when Sharrkan heard that he had a brother (for to that time he knew only of his sister) he turned to his sire and said to him, "O King, hast thou a son other than myself?" He answered, "Yes, and he is now six years old;" adding that his name was Zau al-Makan and that he and Nuzhat al-Zaman were twins, born at a birth. This news was grievous to Sharrkan, but he kept his secret and said, "The blessing of Allah Most High be upon them!"; and he cast the jewel from his hand and shook the dust off his clothes. Quoth the King, "How do I see thee change thy manner when hearing of this, considering that after me thou becomest heir of the kingdom. Of a truth the troops have sworn to thee and the Emirs and Grandees have taken the oath of succession to thee; and this one of the three jewels is thine." Sharrkan bowed his head to the ground and was ashamed to bandy words with his parent so he accepted the jewel and went away, knowing not what to do for exceeding wrath, and stayed not walking till he had entered Abrizah's palace. As he approached she stood up to meet him and thanked him for what he had done and prayed for blessings on him and his sire. Then she sat down and seated him by her side; but when he had taken his place she saw rage in his face and questioned him, whereupon he told her that Allah had blessed his father with two children by Sophia, a boy and a girl, and that he had named the boy Zau al-Makan and the girl Nuzhat al-Zaman; adding, "He hath kept the other two jewels for them and hath given me one of thine, so I left it behind; I knew naught of Zau al-Makan's birth till this day, and the twain are now six years old. So when I learnt this, wrath possessed me; and I tell thee the reason of my rage and hide
- 60. nothing from thee. But now I fear lest my father take thee to wife, for he loveth thee and I saw in him signs of desire for thee: so what wilt thou say, if he wish this?" Quoth she, "Know, O Sharrkan, that thy father hath no dominion over me, nor can he have me without my consent; and if he prevail over me by force, I will take my own life. As for the three jewels, it was not my intent that he should give any of them to either of his children and I had no thought but that he would lay them up in his treasury with his things of price; but now I desire of thy favour that thou make me a present of the jewel which he gave thee, if thou have accepted it." "Hearkening and obedience," replied Sharrkan, and gave it to her. Then said she, "Fear nothing," and talked with him awhile and continued, "I fear lest my father hear that I am with you and sit not patiently under my loss, but do his endeavours to find me; and to that end he may ally himself with King Afridun, on account of his daughter Sophia, and both come on thee with armies and so there befal great turmoil." When Sharrkan heard these words, he said to her, "O my lady, if it please thee to sojourn with us, take no thought of them; though there gather together against us all that be on land and on sea." "'Tis well," rejoined she; "if ye entreat me fair, I will tarry with you, and if ye deal evilly by me, I will depart from you." Then she bade her slave-maidens bring food; so they set the tables, and Sharrkan ate a little and went away to his own house, disturbed and perturbed. Such was his case; but regarding the affairs of his father, Omar bin al-Nu'uman, after dismissing his son Sharrkan he arose and, taking the other two jewels, betook himself to the Lady Sophia, who stood up when she saw him and remained standing till he was seated. Presently, his two children, Zau al-Makan and Nuzhat al- Zaman, came to him and he kissed them and hung a jewel round each one's neck, at which they rejoiced and kissed his hands. Then went they to their mother, who joyed in their joy and wished the King long life; so he asked her, "Why hast thou not informed me all this time that thou art the daughter of King Afridun, Lord of Constantinople, that I might have honoured thee still more and enlarged thee in dignity and raised thy rank?" "O King," answered Sophia, "and what could I desire greater or higher than this my
- 61. standing with thee, overwhelmed as I am with thy favours and thy benefits? And, furthermore, Allah hath blessed me with two children by thee, a son and a daughter." Her reply pleased the King and after leaving her, he set apart for her and her children a wondrous fine palace. Moreover, he appointed for them eunuchs and attendants and doctors of law and doctors of philosophy and astrologers and physicians and surgeons to do them service; and in every way he redoubled his favour and entreated them with the best of treatment. And presently he returned to the palace of his dominion and to his Court where he distributed justice among the lieges. So far concerning him and Sophia and her children; but in the matter of Abrizah the King was greatly occupied with love of her and burnt with desire of her night and day; and every night, he would go in to her and converse with her and pay his court to her, but she gave him no answer, only saying, "O King of the age! I have no desire for men at this present." When he saw her withdraw from him, his passion waxed hotter and his longing and pining increased until, when weary of this, he summoned his Wazir Dandan and, opening his very heart to him, told him of his love for Princess Abrizah, daughter of Hardub, and informed him how she refused to yield to his wishes and how desire for her was doing him to die, for that he could get no grace of her. The Wazir, hearing these words, said to the King, "As soon as it is dark night, take thou a piece of Bhang the measure of a miskal, about an ounce, and go in to her and drink somewhat of wine with her. When the hour of ending the carousal shall draw near, fill her a last cup and dropping therein the Bhang, give it to her to drink, and she will not reach her sleeping-chamber ere the drug take effect on her. Then do thou go in to her and take thy will of her; and such is my advice."[218] "Thy rede is aright," quoth the King, and seeking his treasury, he took thence a piece of concentrated Bhang, if an elephant smelt it he would sleep from year to year. This he put in his bosom-pocket and waited till some little of the night went by, when he betook himself to the palace of Princess Abrizah, who seeing him stood up to receive him; but he bade her sit down. So she sat down, and he sat by her, and he began to talk with her of wine and wassail, whereupon she furnished
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