Laboratory Format for Lab Reports Title Page (Contains ti.docx

Laboratory Format for Lab Reports
Title Page (Contains title, author, course name, experiment
number; and report dates.)
Abstract or Summary: (brief description of experiment's purpose
and outcome)
Hypothesis: (Clearly states the correct purpose/ hypothesis,
which scientific principles are to be
tested and the variables involved.)
Methodology: (In clear, concise sentences with step-by-step
format. Include a list of materials
employed followed by a walkthrough of what you did)
Observations and Results: (pictures followed by a general
description of what you observed,
and/or any relevant data tables created. I love pictures and
illustrations especially of your lab set
up, test tubes before and after, or sketches of in colored pencils
of what you saw under the
microscope. Be sure to include what magnification you are
looking at. I love to see a detailed
observation and description of what you saw. Send the pictures
as individual attachments not
embedded in the report. If the report gets too large, you will not
be able to post it to Blackboard,
so attach each image as a JPEG separately.
Complete description of what occurred stated in complete
sentences. Data is used accurately in
reporting/analyzing the results. Error analysis present and

correct.)
Conclusion or Discussion: (your interpretation of the results as
aided by answering the
questions provided at the end of the lab, and if there was any
deviance from what you think was
supposed to happen) Well written and logical paragraph of
explanation supported by data that
addresses the objectives, scientific principles, and ends with the
‘big picture. Includes scientific
error and pro-poses inquiry for un-answered questions. Should
be at least six sentences.
Answer the Questions: Answer all of the questions from the Lab
Manual from LabPaq. They
provide useful templates that you can just fill in.
References: Include references beyond the book or LabPaq that
enhance your understanding and
interpretation of the laboratory. For online references include
the complete URL or web address
not just “Wikipedia.” I always want to see references !!!
Here is a typical grading form that I use:
Points earned out of 100 possible points:
Format: title, abstract, methods, observation, results,
conclusion, etc. - 20 points
Questions – 30 points
Content – 30 points
References: beyond the text and lab manual – 10 points
Pictures, Sketches, Illustrations – 10 points
Total points earned out of 100 possible points: 100

Good luck to all of you! Enjoy and have fun!
Jim
1
MBK – Lab Report Name: Any Student_______
Section: Lab 1
Observing Bacteria and Blood
September 18, 2010

The purpose of this exercise is to gain knowledge of how
microscopes work
through practical applications of a microscope in the
observation of bacteria and blood.
Hypothesis
This exercise will allow me to gain an understanding of how to
set up and use a
microscope. This will help me gain skills in making
observations about specimens.
Procedures
Time was taken to prepare the microscope for use. The lenses
were wiped
clean. Seven different prepared slides were chosen as
specimens. Two slides were
fresh. See Table 1 below to see which specimens were chosen.
Using a light
microscope, each slide was examined at 10x, 40x, and 100x oil
immersion
magnifications. The observations of each slide were noted. Each
of the slides was
drawn and each drawing was labeled. Magnification was noted
on drawings.
Table 1.

Specimens
“e”
Penicillium with conidia
Bacteria, bacillus form
Yeast
Bacteria, coccus form
Bacteria, spirillium form
Prepared yogurt slide
Fresh yogurt slide - 24 hours old
Fresh Blood smear
3
Observations
The observations are reported in Table 2. Drawings of
specimens follow the
table.
Table 2.
Specimen Magnification Observations
“e” 150x “e” appears upside down
Edges are fuzzy, not clearly defined
Edges of “e” are wavy
Penicillium with conidia 150x Long chains
Flagella-like with bulbous end

600x Fork shaped
Long tail on one end
Bulbous part in the middle
3 to 4 prongs
Small, cube shaped, darker parts are
attached to prong shaped parts
1500x oil immersion Appears to be three dimensional
Brighter colors
2 or 3 Layers of cells
Bacteria, bacillus form 150x Looks more like dirt, than bacteria
Rod shaped
Appear to be touching
600x Very small
Not touching
1500x oil immersion Rod shaped
Dense, not transparent
Assorted lengths
Roughly the same circumference
Yeast 150x Appear to be touching
Pink and blue colored cells
600x Different shades of blue
Overlapping
Many pink cells not touching
Spherical in shape
1500x oil immersion More transparent
Outer membranes are darker
Cells spaced apart from each other
Bacteria, coccus form 150x Cells are in groups
Irregular shapes
Transparent
600x Some cells not in groups
Still difficult to detect shape
1500x oil immersion Short and plump
Not spherical
Dense in color

4
Appear separated
Bacteria, spirillium form 150x Many small groups of cells
Red cells and brown cells
Looks like fibers, the way cells are clustered
Cells look transparent
600x Big spaces between clusters of cells
Rust, yellow, and blue colored cells
Look like boomerangs
1500x oil immersion Some long, some short
Some wavy, some just one bend
transparent
Prepared yogurt slide 150x Very few bacteria in view
Cannot tell their shape
Clustered in groups
600x Cells are more pronounced
Dense in color
Cocci shaped
Streptococci shaped
1500x oil immersion Cocci shaped
Outer membrane looks thicker than center of
cell
Center is more transparent
Fresh yogurt 23 hours
old
150x Gray matter
Not recognizable as cells
600x Differing shades of gray

Very small
Cannot tell shape
There is movement by many cells
1500x oil immersion Some cells look turquoise in color
Irregular shapes
Fresh blood smear 600x Cells are mobile
Hundreds of erythrocytes
Some are stacked together and appear
worm-like
Pale salmon color
Saw one leukocyte
1500x oil immersion Biconcave shape of erythrocyte
Still look worm-like
Erythrocyte has no nucleus
Erythrocyte seems more transparent
Leukocyte iridescent in color
Leukocyte has irregular shape
Leukocyte has nucleus
5
6

It took almost three hours to figure out a good light source for
the microscope
and to figure out how to adjust it to view the slides. That was
because of inexperience.
It was a good learning exercise. It is believed the exercise was
performed correctly and
the expected observations were made.
Conclusions
The exercise was performed thoroughly. The fresh slide
preparation turned out
well.
Questions:
11
A. List the following parts of the microscope and describe the
function of each
The following parts of the microscope are:
a. The eyepiece transmits and magnifies the image from the
objective
lens to your eye.

b. The body (or tube) is connected to the arm of the microscope.
It can
be adjusted up or down to focus the image on the slide, to
change
lenses, or to change slides.
c. The nosepiece is a rotating mount that holds the objective
lenses.
d. The objective lens gathers light from the specimen.
e. The stage is what the specimen rests upon.
f. The condenser is a lens system that aligns and focuses the
light from
the lamp onto the specimen.
g. The illumination mirror allows light to reflect up to the
specimen.
h. The coarse-focus knob is used to bring the specimen into the
focal
plane of the objective lens.
i. The fine-focus knob allows you to make fine adjustments to
focus the
image.
j. The arm is the curved portion that holds the optical parts at a
fixed
distance and aligns them.
k. The clips hold the slide that the specimen is on. It keeps the
slide from
moving.
l. The base supports the weight of all the microscope parts.
B. Define the following microscopy terms:

a. Focus: Focus can be controlled with coarse or fine adjustment
knobs
to make a blurry specimen more well defined.
b. Resolution: Resolution is the capacity of an optical system to
distinguish or separate two adjacent objects or points from one
another.
c. Contrast: Contrast has to do with the difference in lighting
between
adjacent areas of a specimen. The intensity of light can be
changed,
12
the diaphragm can be opened or closed, or chemical stains can
be
applied to the specimen to improve the contrast.
C. Describe your observations from fresh yogurt slide you
prepared in Part III.
Under 10x magnification, the specimen is very gray. It is not
recognizable as
cells. Under 40x magnification, the slide is different shades of
gray. There is
movement. It is difficult to pick out a specific shape. Under
100x oil immersion
magnification, some cells look turquoise in color. The cells are
short and have
irregular shapes.

D. Were there observable differences between your fresh yogurt
slide and the prepared
yogurt slide? If so, describe them.
There was movement in the fresh yogurt slide. Many bacteria
moved in one
direction together. The cells on the fresh yogurt slide had a
turquoise hue to them.
E. Describe the four main bacterial shapes
1. The coccus is a bacterial cell that is spherical, oval, or bean
shaped.
2. The bacillus is a bacterial cell that is cylindrical, or rod like.
It is longer than it
is wide.
3. The spirrilium is a bacterial cell with a spiral shaped
cylinder.
4. The vibros is a bacterial cell that is rod like, that has a slight
curve in it.
F. What are the common arrangements bacteria are found in.
Bacterial can occur in pairs. Paired spheres are called
diplococcic, paired rods
are called diplobacillus.
Bacteria can occur in strands. Spheres linked in chains are
called streptococci.
Rods occurring in chains are called streptobacillus.
Bacteria can occur in clusters. Spheres grouped in clusters are
called
staphylococci.

G. Were you able to identify specific bacterial morphologies on
either yogurt slide? If
so, which types?
They are small and irregularly shaped. They look coccus
shaped on the
prepared and the fresh slides. On the prepared yogurt slide, the
outer
membrane of the cell looks thicker than the center of the cell.
The center of
the cell on the prepared slide is almost transparent.
H. Describe the cells you were able to see in the blood smear.
I saw erythrocytes in the blood smear. They were biconcave.
They had no
nucleus. Some of them appeared stacked together, and looked
wormlike in
shape. Under the microscope they appear salmon colored, not
red. They were
circular in shape.
I saw what appeared to be a leukocyte. It looked more clear
colored and
iridescent. I believe the darker spot in the middle of it is the
nucleus. It was an
irregular shape.
13
I. Are the cells you observed in your blood smear different than
the bacterial cells you

have observed? Why or why not?
The cells in the blood smear are different than the bacterial
cells I observed.
They appear to be bigger than bacterial cells. The red blood
cells cannot
reproduce.
J. What is the purpose of immersion oil? Why does it work?
Immersion oil enhances the resolving power of the microscope.
Oil has the
same optical qualities as glass, and it prevents the peripheral
light passing
through the specimen from scattering into the air.
MicroBiology
LabPaq / Published by: Hands-On Labs, Inc.
[email protected] / www.HOLscience.com / Toll Free
866.206.0773
A laboratory Manual of Small-Scale Experiments
for the independent Study of
Microbiology

LabPaq® is a registered trademark of Hands-On Labs, Inc.
(HOL). The LabPaq referenced in
this manual is produced by Hands-On Labs, Inc. which holds
and reserves all copyrights on
the intellectual properties associated with the LabPaq’s unique
design, assembly, and learning
experiences. The laboratory manual included with a LabPaq is
intended for the sole use by that
LabPaq’s original purchaser and may not be reused without a
LabPaq or by others without the
specific written consent of HOL. No portion of any LabPaq
manual’s materials may be reproduced,
transmitted or distributed to others in any manner, nor may be
downloaded to any public or
privately shared systems or servers without the express written
consent of HOL. No changes may
be made in any LabPaq materials without the express written
consent of HOL. HOL has invested
years of research and development into these materials, reserves
all rights related to them, and
retains the right to impose substantial penalties for any misuse.
Published by: Hands-On Labs, Inc.
3880 S. Windermere St.
Englewood, CO 80110
Phone: Denver Area: 303-679-6252
Toll-free, Long-distance: 866-206-0773
www.HOLscience.com
E-mail: [email protected]
Printed in the United States of America.
The experiments in this manual have been and may be
conducted in a regular formal laboratory
or classroom setting with the users providing their own

equipment and supplies. However, this
manual was especially written for the benefit of the independent
study of students who do not
have convenient access to such facilities. It allows them to
perform college and advanced high
school level experiments at home or elsewhere by using a
LabPaq, a collection of experimental
equipment and supplies specifically packaged to accompany this
manual.
Use of this manual and authorization to perform any of its
experiments is expressly conditioned
upon the user reading, understanding and agreeing to fully abide
by all the safety precautions
contained herein.
Although the author and publisher have exhaustively researched
many sources to ensure
the accuracy and completeness of the information contained in
this manual, we assume no
responsibility for errors, inaccuracies, omissions or any other
inconsistency herein. Any slight of
people, organizations, materials, or products is unintentional.
www.HOLscience.com 2 ©Hands-On Labs, Inc.
50-0230-MB-02
Table of contents
5
ImportantInformationtoHelpStudentswiththeStudyofMicrobiolog
y
Experiments

49 observing Bacteria and Blood
73 Bacterial Morphology
86 AsepticTechnique&CulturingMicrobes
105 IsolationofIndividualColonies
129 DifferentialStaining
141 Methyl red Voges-Proskauer Test
153 AntibioticSensitivity
167 Microbes in the Environment
Appendix
178 PreparationofCultures
181 PreparationofDisinfecting
Solution
183 FinalCleanupInstructions

ImportantInformationtoHelpStudentswith
the Study of Microbiology
Welcome to the study of microbiology. Do not be afraid of
taking this course. By the end of the
semester you will be really proud of yourself and will wonder
why you were ever afraid of the
m-word, microbiology! After their first microbiology class,
most students say they thoroughly
enjoyed it, learned a lot of useful information for their lives,
and only regret not having studied
it sooner.
Microbiology is not some “mystery” science only
comprehendible by eggheads. Microbiology
is simply the study of microscopic living organisms. It will be
easier for you to understand the
world we live in and to make the multitude of personal and
global decisions that affect our
lives and our planet after you have learned about the

characteristics of life around you and
how organisms change and interact with each other, with the
environment, and with you. Plus,
having microbiology credits on your transcript will certainly be
impressive, and your microbiology
knowledge may create some unique job opportunities for you.
This lab manual of microbiology experiments was designed to
accompany any entry level college
or advanced high school level microbiology course. It can be
used by all students, regardless of the
laboratory facilities available to them. Its experiments have
been and continue to be successfully
performed in regular microbiology laboratories. With the
special LabPaq experiments can be
performed at home by independent-study students or at small
learning centers that do not
have formal laboratories. Throughout the manual there are
references about campus-based and
independent study, but all of the information and references
herein are equally relevant to both
types of students.
Introduction

www.HOLscience.com 5 © Hands-On Labs, Inc.
Micro- and Small-Scale Experiments
You may be among the growing number of students to take a
full-credit microbiology course
through independent study. If so, you can thank the
development and perfection of micro-
and small-scale techniques in microbiology experimentation.
Experimentation is essential and
fundamental to fully understanding the concepts of
microbiology. In the past, microbiology
courses required that all classes be conducted on a campus
because experiments had to be
performed in the campus laboratory. This was due in part to the
potential hazards inherent in
some traditional experimentation.
These elements of danger, plus increasing chemical and material
costs and environmental concerns
about chemical and biological material disposal, made high
schools, colleges, and universities
reexamine the traditional laboratory methods used to teach
subjects such as chemistry and

microbiology. Scientists began to scale down the quantities of
chemicals used in their experiments
and found that reaction results remained the same, even when
very tiny amounts of chemicals
were used. Institutions also discovered that student learning was
not impaired by studying small-
sized reactions.
Over time, more and more traditional chemistry and
microbiology experiments were redesigned for
micro- and small-scale techniques. One of the primary pioneers
and most prominent contributors
to micro- and small-scale experimentation is Dr. Hubert Alyea
of Princeton University. He not only
reformatted numerous experiments, he also designed many of
the techniques and equipment
used in micro- and small-scale chemistry and microbiology
today.
With decreased hazards, costs, and disposal problems, micro-
and small-scale experimentation
techniques were quickly adapted for use in scholastic
laboratories. As these techniques continued
to be further refined it became possible to perform basic
experiments in the classroom and

eventually outside the classroom. This slow but steady
progression of micro- and small-scale
techniques makes it possible for independent study students to
take a full-credit microbiology
course since they can now perform experiments at home.
Introduction
HowtoStudyMicrobiology
Microbiology is not the easiest subject to learn, but neither is it
the hardest. As in any other
class, if you responsibly apply yourself, conscientiously read
your text, and thoughtfully complete
your assignments, you will learn the material. Here are some
basic hints for effectively studying
microbiology - or any other subject - either on or off campus.
Plan to Study: You must schedule a specific time and establish a
specific place in which to seriously,
without interruptions or distractions, devote yourself to your
studies. Think of studying like you

would think of a job, except that now your job is to learn. Jobs
have specific times and places in
which to get your work done, and studying should be no
different. Just as television, friends, and
other distractions are not permitted on a job; you should not
permit them to interfere with your
studies. You cannot learn when you are distracted. If you want
to do something well, you must
be serious about it. Only after you’ve finished your studies
should you allow time for distractions.
get in the right Frame of Mind: Think positively about yourself
and what you are doing. Give
yourself a pat on the back for being a serious student and put
yourself in a positive frame of mind
to enjoy what you are about to learn. Then get to work!
Organize any materials and equipment
you will need in advance so you don’t have to interrupt your
thoughts to find them later. Look
over your syllabus and any other instructions to know exactly
what your assignment is and what
you need to do. Review in your mind what you have already
learned. Is there anything that you
aren’t sure about? Write it down as a formal question, then go
back over previous materials to try

to answer it yourself. If you haven’t figured out the answer after
a reasonable amount of time and
effort, move on. The question will develop inside your mind and
the answer will probably present
itself as you continue your studies. If not, at least the question
is already written down so you can
discuss it later with your instructor.
BeActivewiththeMaterial:Learning is reinforced by relevant
activity. When studying feel free
to talk to yourself, scribble notes, draw pictures, pace out a
problem, tap out a formula, etc.
The more active things you do with study materials, the better
you will learn. Have highlighters,
pencils, and note pads handy. Highlight important data, read it
out loud, and make notes. If there
is a concept you are having problems with, stand up and pace
while you think it through. See
the action taking place in your mind. Throughout your day try
to recall things you have learned,
incorporate them into your conversations, and teach them to
friends. These activities will help
to imprint the related information in your brain and move you
from simple knowledge to true
understanding of the subject matter.

Do the Work and Think about What you are Doing: Sure, there
are times when you might get away
with taking a shortcut in your studies, but in doing so you will
probably shortchange yourself. The
things we really learn are the things we discover ourselves. That
is why we don’t learn as much
from simple lectures or when someone gives us the answers.
And when you have an assignment,
don’t just go through the motions. Enjoy your work, think about
what you are doing, be curious,
examine your results, and consider the implications of your
findings. These “critical thinking”
techniques will improve and enrich your learning process. When
you complete your assignments
independently and thoroughly you will have gained knowledge
and you will be proud of yourself.
Introduction
HowtoStudyMicrobiologyIndependently

There is no denying that learning through any method of
independent study is a lot different than
learning through classes held in traditional classrooms. A great
deal of personal motivation and
discipline is needed to succeed in a course of independent study
where there are no instructors or
fellow students to give you structure and feedback. But these
problems are not insurmountable
and meeting the challenges of independent study can provide a
great deal of personal satisfaction.
The key to successful independent study is in having a personal
study plan and the personal
discipline to stick to that plan.
Properly Use your learning Tools: The basic tools for
telecourses, web courses and other
distance-learning methods are often similar and normally
consist of computer software or videos,
textbooks, and study guides. Double check with your course
administrator or syllabus to make sure
you acquire all the materials you will need. These items are
usually obtained from your campus
bookstore, library, or via the Internet. Your area’s public and
educational television channels may
even broadcast course lectures and videos. If you choose to do

your laboratory experimentation
independently, you will need the special equipment and supplies
described in this lab manual
and contained in its companion LabPaq. The LabPaq can be
purchased on the Internet at www.
LabPaq.com.
For each study session, first work through the appropriate
sections of your course materials. These
basically serve as a substitute for classroom lectures and
demonstrations. Take notes as you would
in a regular classroom. Actively work with any computer and/or
text materials, carefully review
your study guide, and complete all related assignments. If you
do not feel confident about the
material covered, repeat these steps until you do. It’s a good
idea to review your previous work
before proceeding to a new section. This reinforces what you
previously learned and prepares you
to absorb new information. Experimentation is the very last
thing done in each study session and
it will only be really meaningful if you have first absorbed the
text materials that it demonstrates.
Plan to Study: A regular microbiology course with a laboratory

component will require you to spend
around 15 hours a week studying and completing your
assignments. Remember, microbiology is
normally a 5-credit hour course! To really learn new material
there is a generally accepted 3-to-
1 rule that states that at least 3 hours of class and study time are
required each week for each
hour of course credit taken. This rule applies equally to
independent study and regular classroom
courses. On campus, microbiology students are in class for 4
hours and in the laboratory for
2 to 3 hours each week. Then they still need at least 8 hours to
read their text and complete
assignments. Knowing approximately how much time you need
will help you to formulate a study
plan at the beginning of the course and then stick with it.
Schedule your Time Wisely: The more often you interact with
study materials and call them to
mind, the more likely you are to reinforce and retain the
information. Thus, it is much better to
study in several short blocks of time rather than in one long,
mind-numbing session. Accordingly,
you should schedule several study periods throughout the week,
or better yet, study a little each

day. Please do not try to do all of your study work on the
weekends! You will just burn yourself
out, you won’t really learn much, and you will probably end up
feeling miserable about yourself
and microbiology. Wise scheduling can prevent such
unpleasantness and frustration.
Introduction
ChoosetheRightPlaceforYourHomeLaboratory
If you are experimenting at home, the best place to perform
your micro- and small-scale
microbiology experiments is in an uncluttered room that has
these important features:
● a door that can be closed to keep out pets and children
● a window or door that can be opened for fresh air ventilation
and fume exhaust
● a source of running water for fire suppression and cleanup

● a counter or table-top work surface
● a heat source such as a stove top, hot dish, or Bunsen burner
The kitchen usually meets all these requirements, but you must
make sure you clean your work
area well both before and after experimentation. This will keep
foodstuff from contaminating your
experiment and your experiment materials from contaminating
your food. Sometimes a bathroom
makes a good laboratory, but it can be rather cramped and
subject to a lot of interruptions.
Review the “Basic Safety” section of this manual to help you
select the best location for your
home-lab and to make sure it is adequately equipped.
Introduction
OrganizationoftheLabManual
Before proceeding with the experiments you need to know what

is expected of you. To find out,
please thoroughly read and understand all the various sections
of this manual.
laboratory Notes: Like all serious scientists you will record
formal notes detailing your activities,
observations, and findings for each experiment. These notes
will reinforce your learning
experiences and knowledge of microbiology. Plus, they will
give your instructional supervisor a
basis for evaluating your work. The “Laboratory Notes” section
of this manual explains exactly
how your lab notes should be organized and prepared.
Required Equipment and Supplies: This manual also contains a
list of the basic equipment
and supplies needed to perform all the experiments. Students
performing these experiments
in a non-lab setting must obtain the “LabPaq” specifically
designed to accompany this manual.
It includes all the equipment, materials, and chemicals needed
to perform these experiments,
except for some items usually found in the average home or
obtainable in local stores. At the
beginning of each experiment there is a “Materials” section that

states exactly which items the
student provides and which items are found in the LabPaq.
Review this list carefully to make sure
you have all these items on hand before you begin the
experiment. It is assumed that campus-
based students will have all the needed equipment and supplies
in their laboratories and that the
instructors will supply required materials and chemicals in the
concentrations indicated.
Laboratory Techniques: While these techniques primarily apply
to full-scale experiments in
formal laboratories, knowledge of them and their related
equipment is helpful to the basic
understanding of microbiology and may also be applicable to
your work with micro- and small-
scale experimentation.
Basic Safety and Micro-scale Safety reinforcement: The use of
this lab manual and the LabPaq,
plus authorization to perform their experiments, are expressly
conditioned upon the user reading,
understanding and agreeing to abide by all the safety rules and
precautions noted. Additional terms
authorizing use of the LabPaq are contained in its purchase

agreement. These safety sections are
relevant to both laboratory and non-laboratory experimentation.
They describe potential hazards
plus the basic safety equipment and safety procedures designed
to avoid such hazards. The Basic
SafetyandMicro-
scaleSafetyReinforcementsectionsarethemostimportantsectionso
fthis
lab manual and should
alwaysbereviewedbeforestartingeachnewexperiment.
Experiments: All experimental materials and procedures are
fully detailed in the laboratory manual
for each experiment. Chemicals and supplies unique for a
specific experiment are contained in a
bag labeled with the experiment number.
Introduction
HowtoPerformanExperiment
Although each experiment is different, the process for

preparing, performing, and recording all
the experiments is essentially the same.
ReviewBasicSafety: Before beginning reread the safety
sections, try to foresee potential hazards,
and take appropriate steps to prevent problems.
ReadthroughtheEntireExperimentbeforeYouStart:Knowing what
you are going to do before
you do it will help you to be more effective and efficient.
OrganizeYourWorkSpace,Equipment,andMaterials: It is hard to
organize your thoughts in
a disorganized environment. Assemble all required equipment
and supplies before you begin
working. These steps will also facilitate safety.
outline your lab Notes: Outline the information needed for your
lab notes and set up required
data tables. This makes it much easier to concentrate on your
experiment. Then simply enter your
observations and results as they occur.
PerformtheExperimentAccordingtoInstructions:Follow exactly
all directions in a step-by-step

format. This is not the time to be creative. DO NOT attempt to
improvise your own procedures!
Think About What you Are Doing: Stop and give yourself time
to reflect on what has happened
in your experiment. What changes occurred? Why? What do
they mean? How do they relate
to the real world? This step can be the most fun and often
creates “light bulb” experiences of
understanding.
CompleteYourLabNotesandAnswerRequiredQuestions:If you
have properly followed all the
above steps, this concluding step will be easy.
clean-up: Blot any minute quantities of unused chemicals with a
paper towel or flush them down
the sink with generous amounts of water. Discard waste in your
normal trash. Always clean your
equipment immediately after use or residue may harden and be
difficult to remove later. Return
equipment and supplies to their proper place, and if working at
home with a LabPaq, store it out
of the reach of children and pets.

Introduction
Antibiotic Sensitivity 24 - 48 hrs. ahead 1 hour 24 - 72 hours 1
hour
EXPEriMENT 11:
Fomite Transmission None 1 - 2 hours 24 - 72 hours Less than 1
hour
EXPEriMENT 12:
Microbes in the Environment None 1 - 3 hours 24 - 72 hours
Less than 1 hour
EXPEriMENT 13: 24 hour intervals
Fungi None Less than 1 hour Up to 1 week 2 - 3 hours
EstimatedTimeRequirementsforEachExperiment
Note: These estimates are provided to help you plan and

schedule your time. They are given per individual
lab performed separately and do not consider time and step
savings possible when several labs are grouped
together. Of course, these are only estimates and your actual
time requirements may differ.
ExperimentNo./Title Preparation Experimenting Incubation
AfterIncubation
EXPEriMENT 1:
Observing Bacteria & Blood None 3 - 4 hours None None
EXPEriMENT 2:
Bacterial Morphology None 3 - 4 hours None None
EXPEriMENT 3:
Aseptic Techniques & Culturing
Microbes None 1 - 2 hours 24 - 48 hours Less than 1 hour
EXPEriMENT 4:
Isolation of Individual Colonies None-use Exp. 3 cultures 3 - 4

hours 24 - 48 hours Less than 1 hour
EXPEriMENT 5: 30 minutes
Differential Staining 24 - 48 hours ahead 3 - 4 hours 24 - 48
hours None
EXPEriMENT 6:
Methyl Red 30 minutes
Voges-Proskauer Test 24 - 48 hours ahead Less than 1 hour 48 -
72 hours 1 hour
Motility Testing 24 - 48 hours ahead Less than 1 hour 24 - 48
hours Less than 1 hour
EXPEriMENT 8:
Carbohydrate 30 minutes
Fermentation Testing 24 - 48 hrs. ahead Less than 1 hour 12 -
24 hours Less than 1 hour

Osmosis 24 - 48 hrs. ahead Less than 1 hour 24 - 72 hours Less
than 1 hour
Introduction
laboratory Notes and lab reports
Normally two basic records are compiled during and from
scientific experimentation activities.
The first record is Lab Notes which you will record as you
perform your actual experiments.
Entries into your lab notebook will be the basis for your second
record, the Lab Report. The Lab
Report formally summarizes the activities and findings of your
experiment and is what is normally
submitted for instructor grading.

Scientists keep track of their experimental procedures and
results through lab notes that are
recorded in a journal-type notebook as they work. In
laboratories these notebooks are often read
by colleagues such as directors and other scientists working on
a project. In some cases scientific
notebooks have become evidence in court cases. Thus, lab notes
must be intelligible to others
and include sufficient information so that the work performed
can be replicated and so there can
be no doubt about the honesty and reliability of the data and of
the researcher.
Notebooks appropriate for data recording are bound and have
numbered pages that cannot be
removed. Entries normally include all of the scientist’s
observations, actions, calculations, and
conclusions related to each experiment. Data is never entered
onto pieces of scratch paper to
later be transferred, but rather is always entered directly into
the notebook. When erroneous data
is recorded, a light diagonal line is drawn neatly through the
error, followed by a brief explanation
as to why the data was voided. Information learned from an
error is also recorded. Mistakes can

often be more useful than successes, and knowledge gained
from them is valuable to future
experimentation.
As in campus-based science laboratories, independent-study
students are normally expected to
keep a complete scientific notebook of their work that may or
may not be periodically reviewed
by their instructor. Paperbound 5x7 notebooks of graph paper
usually work well as science lab
notebooks. Since it is not practical to send complete notebooks
back and forth between instructors
and students for each experiment, independent-study students
usually prepare formal Lab
Reports that are submitted to their instructors along with
regular assignments via e-mail or fax.
Lab notes of experimental observations can be kept in many
ways. Regardless of the procedure
followed, the key question for deciding what kind of notes to
keep is this: “Do I have a clear
enough record so that I could pick up my lab notebook or read
my Lab Report in a few months
and still explain to myself or others exactly what I did?”
Laboratory notes normally include these

components:
Title: This should be the same title stated in the laboratory
manual.
Purpose: Write a brief statement about what the experiment is
designed to
determine or demonstrate.
Procedure: Briefly summarize what you did in performing this
exercise and what
equipment was used. Do not simply copy the procedure
statement from the
lab manual.
Data Tables: Tables are an excellent way to organize your
observational data. Where
Introduction
applicable, the “Procedures” section of the experiment often

advises a table
format for data recording. Always prepare tables before
experimenting so
they will be ready to receive data as it is accumulated.
Observations: What did you observe, smell, hear, or otherwise
measure? Usually,
observations are most easily recorded in table form.
Questions: Questions are asked frequently throughout and at
the end of exercises.
They are designed to help you think critically about the exercise
you just
performed. Answer thoughtfully.
conclusions: What did you learn from the experiment? Your
conclusions should be based
on your observations during the exercise. Conclusions should be
written in
your best formal English, using complete sentences, paragraphs,
and correct
spelling.
Herearesomegeneralrulesforkeepingalabnotebookonyoursciencee
xperiments:

Leave the first two to four pages blank so you can later add a
“Table of Contents” at the front of
the notebook. Entries into the table of contents should include
the experiment number and name
plus the page number where it can be found.
● Your records should be neatly written.
● The notebook should not contain a complete lab report of
your experiment. Rather, it should
simply be a record of what you did, how you did it, and what
your results were. Your records
need to be complete enough so that any reasonably
knowledgeable person familiar with the
subject of your experiment, such as another student or your
instructor, can read the entries,
understand exactly what you did, and if necessary, repeat your
experiment.
● Organize all numerical readings and measurements in
appropriate data tables as in the sample
Lab Report presented later.
● Always identify the units for each set of data you record

(centimeters, kilograms, seconds,
etc.).
● Always iden tify the equipment you are using so you can find
or create it later if needed to
recheck your work.
● It is an excellent idea to document important steps and
observations of your experiments via
digital photos and also to include yourself in these photos. Such
photos within your Lab Report
will document that you actually performed the experiment as
well as what you observed.
● In general, it is better to record more rather than less data.
Even details that may seem to
have little bearing on the experiment you are doing (such as the
time and the temperature
when the data were taken and whether it varied during the
observations) may turn out to be
information that has great bearing on your future analysis of the
results.
● If you have some reason to suspect that a particular data set
may not be reliable (perhaps you

had to make the read ing very hurriedly) make a note of that
fact.
Introduction
● Never erase a reading or data. If you think an entry in your
notes is in error, draw a single line
through it and note the correction, but do not scratch it out
completely or erase it. You may
later find that it was significant after all.
Although experimental results may be in considerable error,
there is never a “wrong” result in
an experi ment for even errors are important results to be
considered. If your observations and
measurements were carefully made, your result will be correct.
Whatever happens in nature,
includ ing the laboratory, cannot be wrong. Errors may have
nothing to do with your investigation,
or they may be mixed up with so many other events you did not
expect that your report is not use-

ful. Yet even errors and mistakes have merit and often lead to
our greatest learning experiences.
Thus, you must think carefully about the interpretation of all
your results, including your errors.
Finally, the cardinal rule in a laboratory is to fully carry out all
phases of your experiments instead
of “dry-labbing” or taking shortcuts. The Greek scientist,
Archytas, summed this up very well in
380 BCE:
In subjects of which one has no knowl edge one must
obtain knowledge either by learning from someone else or
by discover ing it for oneself. That which is learned, there-
fore, comes from another and by outside help; that which is
discovered comes by one’s own efforts and independently.
To discover without seeking is difficult and rare, but if one
seeks it is frequent and easy. If, however, one does not
know how to seek, discovery is im possible.
Introduction

Science lab report Format
This guide covers the overall format that formal Lab Reports
normally follow. Remember that
the Lab Report should be self-contained so that anyone,
including someone without a science
background and without a lab manual, can read it and
understand what was done and what was
learned. Data and calculation tables have been provided for
many of the labs in this manual and
students are encouraged to use them. Computer spreadsheet
programs such as Excel® can greatly
facilitate the preparation of data tables and graphs. One website
with additional information
on preparing lab reports is: http://www.ncsu.edu/labwrite/.
Remember, above average work is
necessary to receive above average grades!
Lab Reports are expected to be word processed and to look
organized and professional. They
should be free of grammar, syntax, and spelling errors and be a
respectable presentation of your
work. Writing in the first person should be avoided as much as
possible. Lab Reports should
generally contain these sections:

● Title Page
● Section 1: Abstract, Experiment Description, Procedures,
and Observations including photos,
drawings, and data tables
● Section 2: Analysis including calculations, graphs, and error
analysis
● Section 3: Discussion of Results
Each of the above three sections is discussed in greater detail
below. They should be clearly
distinguished from each other in the actual report. The
presentation and organization skills
developed by producing science Lab Reports will be beneficial
to all potential career fields.
Title Page: This is the first page of the lab report and consists
of:
a. Experiment number and/or title
b. Your name

c. The names of any lab partner(s)
d. The date and time the experiment was preformed
e. The location should be included if work was performed in the
field
f. The course number
Introduction
Section1:Abstract,Experiment,andObservation
Abstract: Even though the abstract appears at the beginning of
the report, it is written last and
inserted into the beginning. An abstract is a very concise
description of the experiment’s objective,
results, and conclusions. It should be no longer than a
paragraph.
ExperimentandObservation:Carefully, yet concisely, describe,

in chronological order, what was
done, what was observed, and what, if any, problems were
encountered. Describe what field and
laboratory techniques and equipment were employed to collect
and analyze the data upon which
the conclusions are based. Photos and graphic illustrations are
usually inserted in this section.
Graphics should be in .jpg or .gif format to minimize their
electronic file size.
Show all work for any calculations performed. Every graph
must have a title and its axes must be
clearly labeled. Curves through data points this should be “best-
fit curves,” which are smooth
straight or curved lines that best represents the data, rather than
a dot-to-dot connection of data
points.
Include all data tables, photos, graphs, lists, sketches, etc. in an
organized fashion. Include relevant
symbols and units with data. Generally a sentence or two
explaining how data was obtained is
appropriate for each data table.
Note any anomalies observed or difficulties encountered in

collecting data as these may affect the
final results. Include information about any errors observed and
what was learned from them. Be
deliberate in recording the experimental procedures in detail.
Your comments may also include
any preliminary ideas you have on explaining the data or trends
you see emerging.
Introduction
Section2:AnalysisincludingCalculations,Graphs,
and Error Analysis
Generally, the questions at the end of each lab will act as a
guide for preparing results and
conclusions. This section is normally written in paragraph form
and not more than one or two
pages long. Additional considerations are:
● What is the connection between the experimental
measurements taken and the final results
and conclusions? How do your results relate to the real world?

● What were the results of observations and calculations?
● What trends were noticed?
● What is the theory or model behind the experiment
preformed?
● Do the experimental results substantiate or refute the theory?
Why? Be sure to refer
specifically to the results you obtained!
● Were the results consistent with your original predictions of
outcomes or were you forced to
revise your thinking?
● Did “errors” such as environmental changes (wind, rain, etc.)
or unplanned friction occur? If
so, how did they affect the experiment?
● Did any “errors” occur due to the equipment used such as
estimates being skewed due to a
lack of sufficient measurement gradients on a beaker?
● What recommendations might improve the procedures and

results?
Errors: In a single paragraph comment on the accuracy and
precision of the apparatus and include
a discussion of the experimental errors and an estimate of the
error in your final result. Remember,
“errors” are not “mistakes!” Errors arise because the apparatus
and/or the environment inevitably
fail to match the “ideal circumstances” assumed when deriving
a theory or equations. The two
principal sources or error are:
Physical phenomena: Elements in the environment may be
similar to the phenomena
being measured and thus may affect the measured quantity.
Examples might include stray
magnetic or electric fields or unaccounted for friction.
Limitationsoftheobserver,theanalysis,and/ortheinstruments:
Examples are parallax
error when reading a meter tape, the coarse scale of a graph,
and the sensitivity of the
instruments.
Examples of “mistakes” and “human errors” that are not

acceptable scientific errors include:
a. Misuse of calculator (pushing the wrong button, misreading
the display)
b. Misuse of equipment
c. Faulty equipment
d. Incorrectly assembled circuit or apparatus
Introduction
Section3:Discussion,Results,andConclusions
Discussion: The discussion section should be carefully
organized and include consideration of
the experiment’s results, interpretation of results, and
uncertainty in results as further described
below. This section is normally written in paragraph form and
no more than one to two pages
in length. Occasionally it will be more appropriate to organize

various aspects of the discussion
differently for different labs. Not all of the following questions
will apply to every lab.
results
● What is the connection between your observations,
measurements, and final results?
● What were the independent or dependent variables in the
experiment?
● What were the results of your calculations?
● What trends were noticeable?
● How did the independent variables affect the dependent
variables? For example, did an
increase in a given independent variable result in an increase or
decrease in the associated
dependent variable?
InterpretationofResults
● What is the theory or model behind the experiment you

performed?
● Do your experimental results substantiate or agree with the
theory? Why or why not? Be sure
to refer specifically to YOUR experimental results!
● Were these results consistent with your original beliefs or
were you forced to re-evaluate your
prior conceptions?
Uncertainty in results:
● How much did your results deviate from expected values?
● Are the deviations due to error or uncertainty in the
experimental method or are they due to
idealizations inherent in the theory, or are they due to both?
● If the deviations are due to experimental uncertainties can
you think of ways to decrease the
amount of uncertainty?
● If the deviations are due to idealizations in the theory what
factors has the theory neglected
to consider? In either case, consider whether your results

display systematic or random
deviations.
All of these comments on lab notes and lab reports undoubtedly
sound complex and overwhelming
upon first reading. But do not worry; they will make more sense
to you when you actually begin
to perform the experiments and write reports. After writing the
first few lab reports they will
become second nature to you. This manual contains a sample
lab report example of “A” level
work to provide a better understanding of how a formal lab
report is written.
Introduction
LaboratoryDrawings
Laboratory work often requires findings to be illustrated in
representational drawings. Clear,
well organized drawings are an excellent way to convey
observations and are often more easily

understood than long textual descriptions. The adage “a picture
is worth a thousand words” really
is true when referring to science laboratory notes.
Students often say they can’t draw, but with a little care and
practice, anyone can illustrate science
lab observations. A trick most artist’s use is to place a mental
grid over the object or scene and
then approach their drawing from the standpoint of the grid
areas. For instance, look at the
diagram below and quickly make a free hand drawing of it.
Then mentally divide the diagram
into quarters and try drawing it again. In all likelihood, the
second grid-based drawing will yield a
better result.
Give yourself ample drawing space, and leave a white margin
around the actual illustration so it
can be seen clearly. Also, leave a broad margin along one side
of your drawing to insert labels for
the objects in the drawing. Use a ruler to draw straight lines for
the labels and as connecting lines
between the objects and their related labels. The following is a
good example of how your lab
drawings should look when they are included in a formal lab

report.
SoUrcE oF DrAWiNg
Such as MUNG BEAN
your Name
DateofDrawing
TiTlE oF DrAWiNg
Such as CELL STRUCTURE
Introduction
VisualPresentationofData
Learning to produce good graphs and tables is important
because like pictures they can quickly
and clearly communicate information visually. That is why
graphs and tables are often used

to represent or depict data that has been collected. Graphs and
tables should be constructed
in such a way that they are able to “stand alone.” That means,
all the information required to
understand a graph or table must be included in it. A graph is
composed of two basic elements:
the graph itself and the graph legend. The legend adds the
descriptive information needed to
fully understand the graph. In the graph at right the legend
shows that the red line represents
Red Delicious apples, the brown line is the Gala apples, and the
green line is the Wine Sap apples.
Without the legend it would be difficult to interpret this graph.
One of the most important uses of a graph is to “predict” data
that is not measured by the data.
In interpolation a graph is used to construct new data points
within the range of a discrete set of
known data points. As an example, if the data points on the pH
graph are recorded at pHs of 1,
3, 5, 7, 9, and 11 but the investigator wants to know what
happens at pH 6 the information can
be found by interpolating the data between the points of pH 5
and 7. Follow the red line up to
interpolate the value, there would be 12 tadpoles living at a pH

of 6.
Along the same lines, a graph line can be extended to
extrapolate data that is outside of the
measured data. For example, if the researcher wanted to know
what would happen at a pH
greater than 11, this can be extrapolated by extending the line.
In the example at right, the blue
line represents an extrapolation that allows scientists to predict
what might happen. Why is
extrapolation less reliable than interpolation?
Introduction
Concentration of Plant Fertilizer vs. Plant Height
x-axis y-axis
Fertilizer%solution PlantHeightincm
0 25
10 34
20 44

30 76
40 79
50 65
60 40
ConstructingaTable: A table allows for the data to be presented
in a clear and logical way. The
independent data is put at the left hand side of the table and the
dependent data falls to the right
of that. Keep in mind that there will be only one independent
variable but there can be more
than one dependent variable. The decision to present data in a
table rather than a figure is often
arbitrary. However, a table may be more appropriate than a
graph when the data set is too small
to warrant a graph, or it is large and complex and is not easily
illustrated. Frequently, a data table
is provided to display the raw data, while a graph is then used
to make the visualization of the
data easier.
SettingupaGraph:Consider a simple plot of the “Plant Fertilizer”
versus the “Plant Height.”
This is a plot of points on a set of X and Y coordinates. The x-
axis or abscissa, runs horizontally,

while the y-axis or ordinate, runs vertically. By convention, the
x-axis is used for the independent
variable which is defined as a manipulated variable in an
experiment whose presence determines
the change in the dependent variable. The y-axis is used for the
dependent variable which is
the variable affected by another variable or by a certain event.
In our example, the amount of
fertilizer is the independent variable and should go on the x-
axis. The plant height is the dependent
Introduction
variable and should go on the y-axis since it may change as a
result of or dependent on how the
amount of fertilizer changes.
One way to help figure out which data goes on the x-axis versus
the y-axis is to think about what
affects what, so does fertilizer affect plant height or would
plant height affect the fertilizer. Only

one of these should make sense, plant height will not change the
fertilizer but the fertilizer will
have an effect on the plant height. So which ever causes the
change is the independent or x-axis
and which responds as a result of that change is the dependent
or y-axis.
The rules for constructing a table are similar. The important
point is that the data is presented
clearly and logically. As shown in the prior table, the
independent data is put at the left-hand side
of the table and the dependent data falls to the right of that.
Keep in mind that there will be only
one independent variable, but there can be more than one
dependent variable. The decision
to present data in a table rather than a figure is often arbitrary.
However, a table may be more
appropriate than a graph when the data set is too small to
warrant a graph, or it is large and
complex and is not easily illustrated. Frequently, a data table is
provided to display the raw data,
while a graph is then used to make the visualization of the data
easier.
If the data deals with more than one dependent variable such as

the apple varieties seen in the
first example, it would be represented with three lines and a key
or legend would be needed to
identify which line represents which data set. In all graphs each
axis is labeled and the units of
measurement are specified. When a graph is presented in a lab
report, the variables, the scale,
and the range of the measurements should be clear. Graphs are
often the clearest and easiest
way to depict the patterns in your data -- they give the reader a
“feel” for the data.
Use the table below to help set up a line graph. Once you have a
good feel for how to create a
graph on your own, explore computer graphing using MS Excel.
Another easy program to use is
http://nces.ed.gov/nceskids/Graphing/Classic/line.asp
Introduction

Introduction
IntroductiontoMicroscopy
Ever since their invention in the late 1500s, light microscopes
have enhanced our knowledge
of basic microbiology, biomedical research, medical
diagnostics, and materials science. Light
microscopes can magnify objects up to 1000 times, revealing a
world unknown to the naked
eye details. Light-microscopy technology has evolved far
beyond the first microscopes of Robert
Hooke and Antoni van Leeuwenhoek. Special techniques and
optics have been developed to
reveal the structures and biochemistry of living cells.
Microscopes have even entered the digital
age, using fluorescent technology and digital cameras, yet the
basic principles of these advanced
microscopes are a lot like those of the microscope you will use
in this class.
A light microscope works very much like a refracting telescope

but with some minor differences.
Let’s briefly review how a telescope works.
A telescope must gather large amounts of light from a dim,
distant object. Therefore, it needs a
large objectivelens to gather as much light as possible and bring
it to a bright focus. Because the
objective lens is large, it brings the image of the object to a
focus at some distance away which is
why telescopes are much longer than microscopes. The eyepiece
of the telescope then magnifies
that image as it brings it to your eye.
In contrast to a telescope, a microscope must gather light from a
tiny area of a thin, well-
illuminated specimen that is nearby. So the microscope does not
need a large objective lens.
Instead, the objective lens of a microscope is small and
spherical, which means that it has a much
shorter focal length on either side. It brings the image of the
object into focus at a short distance
within the microscope’s tube. The image is then magnified by a
second lens, called an ocular lens
or eyepiece, as it is brought to your eye.

The other major difference between a telescope and a
microscope is that a microscope has a light
source and a condenser. The condenser is a lens system that
focuses the light from the source
onto a tiny, bright spot of the specimen which is the same area
that the objective lens examines.
Also, unlike a telescope, which has a fixed objective lens and
interchangeable eyepieces,
microscopes typically have interchangeable objective lenses and
fixed eyepieces. By changing
the objective lenses (going from relatively flat, low-
magnification objectives to rounder, high-
magnification objectives), a microscope can bring increasingly
smaller areas into view -- light
gathering is not the primary task of the objective lens of a
microscope, as it is with that of a
telescope.
Introduction

The Parts of a light Microscope
A light microscope hAs the following bAsic systems
● Specimen control - to hold and manipulate the specimen
● Stage - where the specimen rests
● clips - to hold the specimen still on the stage.
Because you are looking at a magnified image, even
the smallest movements of the specimen can move
parts of the image out of your field of view
● Illumination - to shed light on the specimen. The simplest
illumination system is a mirror that reflects room light up
through the specimen
● lamp - to produce the light. Typically, lamps are
tungsten-filament light bulbs. For specialized
applications, mercury or xenon lamps may be used to
produce ultraviolet light. Some microscopes even use
lasers to scan the specimen
● condenser - a lens system that aligns and focuses the
light from the lamp onto the specimen

● diaphragm or disc apertures - placed in the light path to alter
the amount of light that
reaches the condenser. Varying the amount of light alters the
contrast in the image
● lenses - to form the image
● objectivelens - to gather light from the specimen
● eyepiece - to transmit and magnify the image from the
objective lens to your eye
● nosepiece - a rotating mount that holds many objective lenses
● tube - to hold the eyepiece at the proper distance from the
objective lens and blocks out
stray light
● Focus - to position the objective lens at the proper distance
from the specimen
● coarse-focus knob - to bring the object into the focal plane of
the objective lens
● fine-focusknob - to make fine adjustments to focus the image

● Support and alignment
● arm - a curved portion that holds all of the optical parts at a
fixed distance and aligns them
● base - supports the weight of all of the microscope parts
● tube - connected to the arm of the microscope by way of a
rack and pinion gear which
allows you to focus the image when changing lenses or
observers and to move the lenses
away from the stage when changing specimens
Introduction
Some of the parts mentioned above may vary between
microscopes. Microscopes come in two
basic configurations: upright and inverted. The microscope
shown in the diagram is an upright
microscope, which has the illumination system below the stage

and the lens system above the
stage. An inverted microscope has the illumination system
above the stage and the lens system
below the stage. Inverted microscopes are better for looking
through thick specimens, such as
dishes of cultured cells, because the lenses can get closer to the
bottom of the dish where the
cells grow.
Light microscopes can reveal the structures of living cells and
tissues as well as of non-living
samples such as rocks and semiconductors. Microscopes can be
simple or complex in design,
and some can do more than one type of microscopy, each of
which reveals slightly different
information. The light microscope has greatly advanced our
biomedical knowledge and continues
to be a powerful tool for scientists
Some Microscope Terms:
● Depthoffield - the vertical distance, from above to below the
focal plane, that yields an
acceptable image

● Fieldofview- the area of the specimen that can be seen
through the microscope with a given
objectivelens
● Focal length - the distance required for a lens to bring the
light to a focus (usually measured
in millimeters)
● Focalpoint/focus - the point at which the light from a lens
comes together
● Magnification - the product of the magnifying powers of the
objective and eyepiece lenses (a
15x eyepiece and a 40x objective lens will give you 15x40=600
power magnification)
● Numerical aperture - the measure of the light-collecting
ability of the lens
● Resolution - the closest two objects can be before they are no
longer detected as separate
objects (usually measured in nanometers)
● ImageQuality - When you look at a specimen using a
microscope, the quality of the image

you see is assessed by the following:
● Brightness - How light or dark is the image? Brightness is
related to the illumination system
and can be changed by changing the wattage of the lamp and by
adjusting the condenser
diaphragm aperture. Brightness is also related to the numerical
aperture of the objective
lens; the larger the numerical aperture, the brighter the image.
● Focus - Is the image blurry or well-defined? Focus is related
to focal length and can be
controlled with the focus knobs. The thickness of the cover
glass on the specimen slide
can also affect your ability to focus the image if it is too thick
for the objective lens. The
correct thickness is usually written on the side of the objective
lens.
Introduction

Image of pollen grain under good brightness (left) and poor
brightness (right)
● Resolution - How close can two points in the image be before
they are no longer seen as
two separate points? Resolution is related to the numerical
aperture of the objective lens
(the higher the numerical aperture, the better the resolution) and
by the wavelength of light
passing through the lens (the shorter the wavelength, the better
the resolution).
Image of pollen grain in focus (left) and out of focus (right)
● contrast - What is the difference in lighting between adjacent
areas of the specimen? Contrast
is related to the illumination system and can be adjusted by
changing the intensity of the
light and the diaphragm/pinhole aperture. Also, chemical stains
applied to the specimen can
enhance contrast.
Image of pollen grain with good resolution (left) and poor
resolution (right)

When observing a specimen by transmitted light, light must pass
through the specimen in order
to form an image. The thicker the specimen the less light passes
through and thereby the darker
the image. The specimens must therefore be thin (0.1 to 0.5
mm). Many organic specimens must
be cut into thin sections before observation. Specimens of rock
or semiconductors are too thick
Introduction
to be sectioned and observed by transmitted light, so they are
observed by the light reflected
from their surfaces.
Image of pollen grain with good contrast (left) and poor
contrast (right)
Introduction

Types of Microscopy
A major problem in observing specimens under a microscope is
that their images do not have
much contrast. This is especially true of living things (such as
cells), although natural pigments,
such as the green in leaves, can provide good contrast. One way
to improve contrast is to treat
the specimen with colored pigments or dyes that bind to specific
structures within the specimen.
Different types of microscopy have been developed to improve
the contrast in specimens. The
specializations are mainly in the illumination systems and the
types of light passed through the
specimen. Brightfield is the basic microscope configuration (the
images seen thus far are all
from brightfield microscopes). This technique has very little
contrast and much of the contrast is
provided by staining the specimens. A darkfield microscope
uses a special condenser to block out
most of the bright light and illuminate the specimen with
oblique light, much like the moon blocks
the light from the sun in a solar eclipse. This optical set-up

provides a totally dark background and
enhances the contrast of the image to bring out fine details of
bright areas at boundaries within
the specimen.
Following are various types of light microscopy techniques.
They achieve different results by using
different optical components. The basic idea involves splitting
the light beam into two pathways
that illuminate the specimen. Light waves that pass through
dense structures within the specimen
slow down compared to those that pass through less dense
structures. As all of the light waves
are collected and transmitted to the eyepiece, they are
recombined, so they interfere with each
other. The interference patterns provide contrast. They may
show dark areas (more dense) on a
light background (less dense), or create a type of false three-
dimensional (3D) image.
● Phase-contrast – A phase-contrast microscope is best for
looking at living specimens, such
as cultured cells. The annular rings in the objective lens and the
condenser separate the light
paths. Light passing through the central part of the light path is

then recombined with light
traveling around the periphery of the specimen. Interference
produced by these two paths
produces images in which dense structures appear darker than
the background.
A phase-contrast image of a glial cell cultured from a rat brain
● Differential InterferenceContrast (DIC) - DIC uses polarizing
filters and prisms to separate
and recombine the light paths, giving a 3D appearance to the
specimen (DIC is also called
Nomarski after the man who invented it).
● HoffmanModulationContrast - Hoffman modulation contrast
is similar to DIC except that it
uses plates with small slits in both the axis and the off-axis of
the light path to produce two
sets of light waves passing through the specimen. Again, a 3D
image is formed.
Introduction

● Polarization - The polarized-light microscope uses two
polarizers, one on either side of the
specimen, positioned perpendicular to each other so that only
light that passes through the
specimen reaches the eyepiece. Light is polarized in one plane
as it passes through the first
filter and reaches the specimen. Regularly-spaced, patterned or
crystalline portions of the
specimen rotate the light that passes through. Some of this
rotated light passes through
the second polarizing filter, so these regularly spaced areas
show up bright against a black
background.
● Fluorescence - This type of microscope uses high-energy,
short-wavelength light (usually
ultraviolet) to excite electrons within certain molecules inside a
specimen, causing those
electrons to shift to higher orbits. When they fall back to their
original energy levels, they
emit lower-energy, longer-wavelength light (usually in the
visible spectrum), which forms the
image.

CareandHandlingoftheMicroscope
● When you move your microscope, you should always use two
hands. Place one hand around
the arm, lift the scope, and put your other hand under the base
of the scope for support. If
you learn to carry the scope in this way, it will force you to
carry it carefully, ensuring that you
do not knock it against anything while moving from one place
to another.
● When you put the scope down, do so gently. If you bang your
scope down on the table
eventually you could jar lenses and other parts loose. Your
microscope seems like a simple
instrument but each eyepiece and objective is actually made up
of a number of lenses put
together in a precise way to create wonderful magnification. If
you bang your scope around,
you are shaking upward of 15 to 20 lenses.
● Always have clean hands when handling your scope. It would
be a shame to damage your
scope with too much peanut butter!

Introduction
Storing the Microscope
● If you have a sturdy, stable desk, table, or shelf on which to
keep your scope and it is a place
where the scope will not be disturbed or bumped, this is the best
place to store your scope.
Just make sure that you keep it covered with a plastic or vinyl
cover when it is not in use. Dust
is an enemy to your lenses; always keep your scope covered
when not in use.
● If you are unable to find a safe place where you can leave
your scope out, store it in the fitted
foam case it comes in.
cleaning the Microscope
● The first step in keeping your microscope clean is to keep it

from getting dirty. Always keep
your microscope covered with the dust cover when it is not in
use.
● Your eyepiece will need cleaning from time to time. Due to
its position on the scope, it will
have a tendency to collect dust and even oil from your
eyelashes. The eyepiece lens should be
cleaned with a high-quality lens paper, such as is available from
a camera shop or an eyeglass
center. Brush any visible dust from the lens and then wipe the
lens. You may wish to use a
bit of lens solution, applied to the lens paper to aid in cleaning.
A cotton swab can be used in
place of lens paper but do not use facial tissues to clean your
lenses.
● You will also need to occasionally clean the objective lenses.
Use a fresh area of lens paper
each time so that you don’t transfer dust from one lens to
another.
● Clean the lenses in the glass condenser under the stage.
● Finally, clean the glass lens over your light, or the mirror, so

that an optimal amount of light
can shine through. You can also follow up by wiping down the
whole scope with a soft, clean
cotton towel.
Using the Microscope
● Take the microscope body from the case. Put the eyepiece in
the opening in the tube at the
top of the microscope. Remove the objective lenses from their
individual containers and screw
them into the revolving nosepiece, placing each in the color-
coded position that corresponds
to the color band on the lens.
● Adjust the tension on the focusing control knobs to suit your
touch or to compensate for
normal wear over time. To increase tension, hold the right-hand
knob firmly and turn the
opposite knob clockwise, whereas turning it counter clockwise
loosens the tension.
● Unplug the rotating mirror bracket from the base of the
microscope, insert the mirror
(packaged separately with the microscope) into the bracket so

that it swivels freely, and plug
it back into the base of the microscope.
● Tilt the arm of the microscope back until it is at a position
where you can comfortably look
into the microscope eyepiece.
Introduction
● Place a slide under the clips on the stage, with the area you
wish to view in line, between the
lens selected and the hole in the stage.
● Turn the nosepiece of the microscope to select the longest
lens (usually the highest power
lens). Lower the barrel of the microscope with the coarse-focus
knob until it almost touches
the slide. If it will not go that far then unscrew the focus stop
screw under the arm of the
microscope until the lens can almost touch the slide, and while
it is in that position lightly

tighten the screw and lock it in place with the knurled nut.
● Place a light source in front of the microscope, use the small
lever on the sub-stage condenser
to open the diaphragm fully, and adjust the mirror so that the
light is brightest as seen through
the microscope.
● Rotate the nosepiece to select the lowest power lens. Lower
the barrel with the coarse-focus
knob until the tip of the lens is near the slide. Now raise the
barrel slowly with the coarse-
focus knob until you see an image from the slide. Finish the
focus with the fine-focus knob.
● With thumb and forefinger on each end of the slide, move it
slowly on the stage until the
object you wish to study is centered in your field of view.
● Rotate the nosepiece of the microscope to select the objective
lens that will give you the
higher magnification you need.
● Once one lens is focused properly any other objective lens on
the nosepiece rotated into

position will be roughly in focus, requiring only fine focus to
bring the image with the new
lens into correct focus.
● Move the lever for the diaphragm through its full range to
select the amount of light that gives
you the best contrast. Many details will be visible with good
contrast which would otherwise
be lost with much or too little light.
Using the Electric illuminator
Grasp the illuminating mirror with your fingers behind its
bracket and pull to unplug the bracket
and mirror from the base of the microscope. Insert the metal
plug tip of the electric illuminator
into the hole from which you have unplugged the mirror
bracket. Rotate the fixture so that the
glass opening over the bulb points up toward the light
condenser under the stage. Plug the electric
cord into a 115-volt outlet and turn on the switch in the cord.
UsingtheOilImmersionLens(purchasedseparately)
Install the oil immersion 100x objective lens in place of any of

the other objective lenses. The 4x
lens is a good choice. First, focus the microscope and center the
slide using a lower magnification
objective. Apply a drop of oil on the specimen slide and turn the
revolving nosepiece to bring the
100x objective into position. If the barrel is too low to allow the
100x lens to move into position
raise it with the coarse focus very slightly, position the lens,
and then lower the barrel until the tip
of the 100x lens touches the oil and the slide. The tip of the lens
is able to move a short distance
into the lens against a spring in order to keep from putting too
much pressure on the slide. With
the lens tip touching the oil and slide focus with the fine-focus
knob. The working distance of the
lens is very short so do not use the coarse-focus knob other than
to position the lens. After using
the oil immersion lens wipe off the oil carefully with alcohol.
Introduction

PreparationofSolidMedia
Microbiological media are used to grow microbes for study and
experimentation. Most bacteria
collected in the environment will not be harmful. However,
once an isolated microbe multiplies
by millions in a broth tube or Petri dish it can become more of a
hazard. Be sure to protect
open cuts with rubber gloves and never ingest or breathe in
growing bacteria. Keep growing Petri
dishes taped closed until your experiment is done. Then you
should safely destroy the bacteria
colonies using bleach.
Microbiological media may be prepared as either liquid or as a
solid media. When a solid medium
is prepared, a corresponding liquid broth is solidified by the
addition of agar to the broth. Agar
is a polysaccharide found in the cell walls of some algae. It is
inert and degraded by very few
microorganisms. In addition, the fact it melts at around 100oC
and solidifies at approximately
45oC-50oC makes it an ideal solidifying agent for
microbiological media.
proceDUres

PreparationofSolidMedia
1. Disinfect your work area with a 10% bleach solution.
2. Place the test tube rack into a pan of water and place your
tubes of agar into the rack. The
agar will melt more easily if the water level is above or at the
level of the agar. If your pan
is not deep enough to bring the water above the level of the agar
you will need to shake the
tubes during the melting process to mix the melted and
unmelted portion of the agar.
3. Place the pan on the stove top and bring to a boil. Once the
water begins to boil, the agar
should melt within 10 to 15 minutes. Remember, if your water
level is below the level of the
agar you will need to shake the tubes to mix the unmelted agar
into the melting agar. Be
careful as the heating tubes will be hot!
4. Once the agar media is melted, remove the pan from the heat
but do not remove the tubes
from the hot water.

5. Allow the water to cool until the tubes are cool enough to
handle but the agar media is still
liquid (50°C-60°C).
6. Label the bottom of two Petri dishes (per tube) with the type
of medium you are using (in this
LabPaq you will use nutrient or MRS agar).
7. Using aseptic handling techniques pour the liquid agar from
the 18-mL tube into the bottom
of the labeled Petri dishes. If you are preparing both types of
medium, be careful to pour each
medium into the correctly labeled dish. Pour enough to cover
the bottom of each dish 1/8”-
1/4” thick (approximately 9mL so each 18-mL tube will make
two dishes). Cover each dish
with its lid immediately.
8. When all the dishes are poured, cover them with a paper
towel to help prevent contamination
and allow them to cool and solidify.
9. The agar dishes are done when solid. You may store the
cooled dishes in a zip baggie in the
refrigerator for later use or use them immediately.

Introduction
PreparationofCultures
Culturetubesshouldremainliddedwhileincubating.Donotopenthe
monceinoculated
unlessunderasepticconditionsandtoperformanecessaryexperiment
alstep.
Saccharomyces cervicae: Add 1/2 teaspoon dry Saccharomyces
cervicae (active dry yeast
envelope) to 1/8 cup warm water (you can use a sample cup or
any household cup) and gently
swirl to mix. Set the culture aside to activate for at least 10
minutes. Stir to mix prior to using.
Escherichia coli:
10. Remove the tube labeled: Broth, Nutrient - 5 mL in Glass
tube, from culture media bag #2
from the refrigerator and allow it to come to room temperature..
11. Moisten a paper towel with a small amount of alcohol and

wipe the work area down.
12. Once the nutrient broth media is at room temperature:
● Remove the numbered E-coli culture tube from the cultures
bag and remove its cap. Set
the cap upside down to avoid contamination.
● Uncap the nutrient broth; set its cap upside down to avoid
contaminating it while the
broth is open.
● Use sterile techniques and draw 0.25mL of the nutrient broth
into a sterile graduated
pipet. NOTE: To sterilize the pipet draw a small amount of 70%
alcohol into the bulb,
and then expel it into a sink. Remove any excess alcohol by
forcefully swinging the pipet
in a downward arch several times to ensure that the pipet is dry
before drawing up the
nutrient broth. Add the broth to the vial containing the
lyophilized E-coli pellet. Recap the
E-coli vial and shake to mix until the pellet has dissolved in the
broth. Note that the vial
should be about one-half full to allow for shaking and mixing
the pellet.

● Once the pellet has dissolved, use the same sterile pipet to
draw up the E. coli solution
and expel it into the original tube of nutrient broth. Recap the
broth. NOTE: If the pipet
has become contaminated, simply draw a small amount of 70%
alcohol into the bulb and
then expel it into a sink. Remove any excess alcohol by
forcefully swinging the pipet in a
downward arch several times to ensure that the pipet is dry
before drawing up the E. coli
solution.
Recap the nutrient broth and incubate the now E-coli inoculated
tube of nutrient broth at 37°C.
The culture should show active growth between 24 to 48 hours;
it can be left as a liquid culture
or plated out. Most freeze dried cultures will grow within a few
days however some may exhibit
a prolonged lag period and should be given twice the normal
incubation period before discarding
as non-viable.
Refer to Experiment 3 for a description of indicators of growth.
● Lactobacillus acidophilus: Remove a tube of MRS broth from

the refrigerator and allow it to
come to room temperature. Aseptically transfer a portion of a
tablet of L. acidophilus into the
tube of media. Allow the tube to set, swirling periodically, as
the tablet dissolves. There will
be a significant amount of sediment in the bottom of the tube.
Mark the level of the sediment
with a marker, pencil, or pen. Incubate the inoculated tube at
37°C. The culture should show
active growth between 24 to 48 hours. Refer to Experiment 3 for
a description of indicators
of growth. L. acidophilus often sediments as it grows. An
increase (above the sediment line
you marked on the tube) in the sediment is an indication of
growth. Swirl the tube to mix the
Introduction
organisms back into the broth prior to use.
● Staphylococcus epidermidis: You can culture S. epidermidis
as a liquid or solid culture.

Because you are inoculating from an environmental source
(your skin), your sample may
contain bacteria other than S. epidermidis. Thus, broth cultures
derived directly from sampling
may not be pure cultures of S. epidermidis. With the exception
of Experiments 3 and 4 (#3
establishes a broth culture and #4 uses it to establish a pure
culture), use the dish culture
method to ensure you are using a pure sample for your
experiment.
● BrothculturesofS.epidermidis: Without contaminating the
cotton tip, cut the length of the
swab such that it will fit entirely into a capped test tube.
Dampen the cotton tip sterile swab
with distilled water and rub it vigorously on your skin. Do not
try to obtain a bacterial culture
soon after washing your skin. Additionally choose an area that
is not as likely to have been
scrubbed as recently (the inside of the elbow or back of the
knee is generally a good site).
Donotobtainasamplefromanybodilyorifice(mouth,nose,etc.)asyo
uarenotlikelyto
culturethedesiredmicrobe(Staphylococcus epidermidis).Using

aseptic technique, place the
swab into a tube of nutrient media, label the tube accordingly.
Incubate the inoculated tube
at 37°C. The culture should show active growth between 24 to
48 hours. Refer to Experiment
3 for a description of indicators of growth.
● DishculturesofS.epidermidis: Use a sterile swab to obtain a
sample of S. epidermidis from
your skin described in the generation of a broth culture. Rub the
swab lightly on the surface
of one dish of nutrient agar to inoculate it with S. epidermidis.
As the swab may not contain
a high number of bacteria, be sure to rub all sides of the swab
on the dish to transfer as
many individual bacterium as possible. Incubate the dish at
37°C for 24 to 48 hours. The S.
epidermidis culture was not a pure culture (derived from a
single organism) and will most
likely contain colonies from several different organisms. You
will need to identify and select
a colony. Staphylococci produce round, raised, opaque colonies,
1 – 2 mm in diameter. S.
epidermidis colonies are white in color. Below is a picture of S.
epidermidis grown on blood agar.

As the sample is of human origin, it potentially contains
bacteria that can act as opportunistic
pathogens. Do not select or use any colony that does not appear
to be S. epidermidis. If your
dish contains colonies other than S. epidermidis, soak it in a
10% bleach solution and discard.
Do not attempt to save the dish for use in future experiments!
You can either use the S. epidermidis colonies directly or
amplify growth in a broth culture. If you
choose to amplify into nutrient broth, 24 hours beginning the
experiment, choose a S. epidermidis
colony from the incubated dish and aseptically transfer the
colony using an inoculation loop into a
tube of nutrient media. Be sure to mix the broth gently to
disburse the clumped bacteria into the
broth. Incubate the tube at 37°C for an additional 24 hours.
Introduction
Microbiology Safety

● Anymicrobecanbehazardous.While the majority of
microorganisms are not pathogenic
to humans and have never been shown to cause illness, under
unusual circumstances a few
microorganisms that are not normally pathogenic can act as
pathogens. These are called
opportunistic pathogens. Treat all microorganisms—especially
unknown cultures such as
from skin swabs or environmental samples—as if they were
pathogenic. A student who
has a compromised immune system or has had a recent extended
illness is at higher risk
for opportunistic infections.
Donotattempttoswabyourthroatornasalpassageswhen
sampling for S. epidermidis. You are not likely to culture the
correct organism. Additionally,
you are more likely to culture an opportunistic pathogen from
these areas!
● Sterilizeequipmentandmaterials. All materials, media, tubes,
dishes, loops, needles, pipets,
and other items used for culturing microorganisms should be
sterilized. Most of the materials
and media you will be using are commercially sterilized

products. You will be given instruction
for sterilization with either flame or with a 10% bleach solution
for items that are not sterilized
or that will be reused.
● Disinfectworkareasbeforeandafteruse.Use a disinfectant, such
as a 10% bleach solution
to wipe down benches and work areas both before and after
working with cultures. Also be
aware of the possible dangers of the disinfectant. Bleach, if
spilled, can ruin your clothing and
can be dangerous if splashed into the eyes. Students should
work where a sink is located to
facilitate immediate rinsing if bleach is splashed or spilled.
● Washyourhands.Use an antibacterial soap to wash your hands
before and after working
with microorganisms. Non-antibacterial soap will remove
surface bacteria and can be used if
antibacterial soap is not available. Gloves should be worn as an
extra protection.
● Neverpipetbymouth. Use pipet bulbs or pipet devices for the
aspiration and dispensing of
liquid cultures.

● Donoteatordrinkwhileworkingwithmicroorganisms.Never eat
or drink while working
with microorganisms. Keep your fingers out of your mouth, and
wash your hands before and
after the laboratory activity. Cover any cuts on your hands with
a bandage. Gloves should be
worn as an extra protection.
● Labeleverythingclearly. All cultures, chemicals,
disinfectants, and media should be clearly
and securely labeled with their names and dates.
● Disinfectallwastematerial. All items to be discarded after an
experiment, such as culture
tubes, culture dishes, swabs, and gloves, should be covered with
a 10% bleach solution
and allowed to soak for at least 1 to 2 hours. After soaking, the
materials can be rinsed and
disposed of by regular means.
● Cleanupspillswithcare.Cover any spills or broken culture
tubes with a 10% bleach solution;
then cover with paper towels. After allowing the spill to sit with
the disinfectant, carefully

clean up and place the materials in a bag for disposal. If you are
cleaning up broken glass, place
the materials in a puncture-proof container (such as a milk
carton), and label the container
“broken glass” before placing in the trash. Wash the area again
with disinfectant. Never pick
Introduction
up glass fragments with your fingers or stick your fingers into
the culture itself. Instead, use a
brush and dustpan.
● Becertaintodisposeofculturesproperly. Liquid cultures should
have bleach added to them
(to create a solution that is approximately 10% bleach) and
allowed to set for a minimum
of one hour before disposal. The deactivated samples can be
discarded in the sink. Be sure
to flush with plenty of water to remove any bleach residue. Petri
dishes or any solid culture

material should be soaked in a 10% bleach solution for a
minimum of one hour. They can then
be bagged and discarded in the trash.
Introduction
Basic Safety guidelines
This section contains vital information that must be thoroughly
read and completely understood
before a student begins to perform experiments.
PrEVENT iNJUriES AND AcciDENTS! Science
experimentation is fun, but does involve potential
hazards which must be acknowledged to be avoided. To safely
conduct science experiments,
students must first learn and then always follow basic safety
procedures. Although there are
certainly not as many safety hazards in experimenting with
physics and geology as there are in
chemistry and biology, safety risks exist in all science
experimentation and science students need

to be aware of safety issues relevant to all the disciplines. Thus,
the following safety procedures
review is relevant to all students regardless of their field of
study.
While this manual tries to include all relevant safety issues, not
every potential danger can be
foreseen as each experiment involves slightly different safety
considerations. Thus, students
must always act responsibly, learn to recognize potential
dangers, and always take appropriate
precautions. Regardless of whether a student will be working in
a campus or home laboratory
setting, it is extremely important that he or she knows how to
anticipate and avoid possible
hazards and to be safety conscious at all times.
BASic SAFETy ProcEDUrES: Science experimentation often
involves using toxic chemicals,
flammable substances, breakable items, and other potentially
dangerous materials and
equipment. All of these things can cause injury and even death
if not properly handled. These
basic safety procedures apply when working in a campus or
home laboratory.

● Because eyesight is precious and eyes are vulnerable to
chemical spills and splashes, to
shattered rocks and glass, and to floating and flying objects:
» Students must always wear eye protecting safety goggles
when experimenting
● Because toxic chemicals and foreign matter may enter the
body through digestion:
» Drinking and eating are always forbidden in laboratory areas
» Students must always wash their hands before leaving their
laboratory
» Students must always clean their lab area after
experimentation
● Because toxic substances may enter the body through the
skin and lungs:
» The laboratory area must always have adequate ventilation
» Students must never “directly” inhale chemicals

» Students should wear long-sleeved shirts, pants, and enclosed
shoes when in their lab
area
» Students must wear gloves and aprons when appropriate
● Because hair, clothing, and jewelry can create hazards, cause
spills, and catch fire while
experimenting:
» Students should always tie or pin back long hair
Introduction
» Students should always wear snug fitting clothing (preferably
old)
» Students should never wear dangling jewelry or objects
● Because a laboratory area contains various fire hazards:

» Smoking is always forbidden in laboratory areas
● Because chemical experimentation involves numerous
potential hazards:
» Studentsmustknowhowtolocateandusebasicsafetyequipment
» Studentsmustneverleaveaburningflameorreactionunattended
» Studentsmustspecificallyfollowallsafetyinstructions
» Students must never perform any unauthorized experiments
»
Studentsmustalwaysproperlystoreequipmentandsuppliesandensur
etheseareout
of the reach of small children and pets
● Because science equipment and supplies often include
breakable glass and sharp items that
pose potential risks for cuts and scratches and small items as
well as dangerous chemicals that
could cause death or injury if consumed:

» Students must carefully handle all science equipment and
supplies
» Students must keep science equipment and supplies stored out
of the reach of pets and
small children
» Students must ensure pets and small children will not enter
their lab area while they are
experimenting
● Because science experimentation may require students to
climb, push, pull, spin, and whirl:
» Students should undertake these activities cautiously and
with consideration for people,
property, and objects that could be impacted
» Students must ensure any stool, chair, or ladder used to climb
is sturdy and take ample
precautions to prevent falls
● Because students’ best safety tools are their own minds and
intellectual ability:

» Students must always preview each experiment, and carefully
think about what safety
precautions need to be taken to perform the experiment safely
BASIC SAFETY EQUIPMENT: The following pieces of basic
safety equipment are found in all
campus laboratories. Informal and home laboratories may not
have or need all of these items,
but simple substitutes can usually be made or found. Students
should know their exact location
and proper use.
SAFETy gogglES - There is no substitute for this important
piece of safety equipment! Spills and
splashes do occur, and eyes can very easily be damaged if they
come in contact with laboratory
chemicals, shattered glass, swinging objects, and flying rock
chips. While normal eyeglasses
do provide some protection, these items can still enter the eyes
from the side. Safety goggles
Introduction

cup around all sides of the eyes to provide the most protection
and can be worn over normal
eyeglasses if required.
EYEWASHSTATION-All laboratories should have safety
equipment to wash chemicals from the
eyes. A formal eyewash station looks like a water fountain with
two faucets directed up at spaces
to match the space between the eyes. In case of an accident, the
victim’s head is placed between
the faucets while the eyelids are held open so the faucets can
flush water into the eye sockets and
wash away the chemicals. In an informal laboratory, a hand-
held shower wand can be substituted
for an eyewash station. After the eyes are thoroughly washed, a
physician should be consulted
promptly.
FIREEXTINGUISHER-There are several types of fire
extinguishers, at least one of which should be
found in all types of laboratories. Students should familiarize
themselves with and know how to
use the particular type of fire extinguisher in their laboratory.

At a minimum, home laboratories
should have a bucket of water and a large pot of sand or dirt
available to smother fires.
FirE BlANKET - This is a tightly woven fabric used to smother
and extinguish a fire. It can cover a
fire area or be wrapped around a victim who has caught on fire.
SAFETYSHOWER-This shower is used in formal laboratories to
put out fires or douse people
who have caught on fire or suffered a large chemical spill. A
hand-held shower wand is the best
substituted for a safety shower in a home laboratory.
FirST-AiD KiT - This kit of basic first-aid supplies is used for
the emergency treatment of injuries
and should be found in both formal and informal laboratories. It
should be always well stocked
and easily accessible.
SPill coNTAiNMENT KiT - This kit consists of absorbent
material that can be ringed around a
spilled chemical to keep it contained until the spill can be
neutralized. The kit may simply be a
bucket full of sand or other absorbent material such as kitty

litter.
FUMEHOOD-This is a hooded area containing an exhaust fan
that expels noxious fumes from
the laboratory. Experiments that might produce dangerous or
unpleasant vapors are conducted
under this hood. In an informal laboratory such experiments
should be conducted only with
ample ventilation and near open windows or doors. If a kitchen
is used for a home laboratory, the
exhaust fan above the stove substitutes nicely for a fume hood.
POTENTIAL LABORATORYHAZARDS: Recognizing and
respecting potential hazards is the first
step toward preventing accidents. Please appreciate the grave
dangers the following laboratory
hazards represent. Work to avoid these dangers and consider
how to respond properly in the
event of an accident.
FirES: The open flame of a Bunsen burner or any heating source
combined, even momentarily,
with inattention may result in a loose sleeve, loose hair, or
some unnoticed item catching fire.
Except for water, most solvents including toluene, alcohols,

acetones, ethers, and acetates which
are highly flammable and should never be used near an open
flame. As a general rule NEVER
LEAVE AN OPEN FLAME OR REACTION UNATTENDED. In
case of fire, use a fire extinguisher, fire
blanket and/or safety shower.
CHEMICALSPILLS:Flesh burns may result if acids, bases, or
other caustic chemicals are spilled and
Introduction
come in contact with skin. Flush the exposed skin with a gentle
flow of water for several minutes
at a sink or safety shower. Acid spills should be neutralized
with simple baking soda, sodium
bicarbonate. If eye contact is involved use the eyewash station
or its substitute. Use the spill
containment kit until the spill is neutralized. To better protect
the body from chemical spills, wear
long-sleeved shirts, full-length pants, and enclosed shoes, not

sandals, when in the laboratory.
AciD SPlATTEr: When water is added to concentrated acid the
solution becomes very hot and
may splatter acid on the user. Splattering is less likely to occur
if acid is slowly added to the water:
Remember this AAA rule: Always Add Acid to water, NEVER
add water to acid.
GLASSTUBINGHAZARDS:Never force a piece of glass tubing
into a stopper hole. The glass may
snap and the jagged edges can cause a serious cut. Before
inserting glass tubing into a rubber or
cork stopper hole be sure the hole is the proper size. Lubricate
the end of the glass tubing with
glycerol or soap, and then while grasping it with a heavy glove
or towel, gently but firmly twist the
tubing into the hole. Treat any cuts with appropriate first-aid.
HEATEDTESTTUBESPLATTER:Splattering and eruptions can
occur when solutions are heated
in a test tube. Thus, you should never point a heated test tube
toward anyone. To minimize this
danger direct the flame toward the top, rather than the bottom,
of the solution in a test tube.

Gently agitate the tube over the flame to heat the contents
evenly.
SHATTERED GLASSWARE: Graduated cylinders, volumetric
flasks and certain other pieces of
glassware are NOT designed to be heated. If heated, they are
likely to shatter and cause injuries.
Always ensure you are using heatproof glass before applying it
to a heat source. Special caution
should always be taken when working with any type of
laboratory glassware.
INHALATIONOFFUMES: To avoid inhaling dangerous fumes,
partially fill your lungs with air and,
while standing slightly back from the fumes, use your hand to
waft the odors gently toward your
nose and then lightly sniff the fumes in a controlled fashion.
NEVER INHALE FUMES DIRECTLY!
Treat inhalation problems with fresh air and consult a physician
if the problem appears serious.
INGESTIONOFCHEMICALS:Virtually all the chemicals found
in a laboratory are potentially toxic.
To avoid ingesting dangerous chemicals, never taste, eat, or
drink anything while in the laboratory.

All laboratories, especially those in home kitchens, should
always be thoroughly cleaned after
experimentation to avoid this hazard. In the event of any
chemical ingestion immediately consult
a physician.
HORSEPLAY:A laboratory full of potentially dangerous
chemicals and equipment is a place for
serious work, not for horseplay! Fooling around in the
laboratory is just an invitation for an
accident.
VEry iMPorTANT cAUTioN For WoMEN: If you are pregnant
or could be pregnant, you should
seek advice from your personal physician before doing any type
of science experimentation.
If you or anyone accidentally consumes or otherwise comes into
contact with something that is
not easily washed away (such as splashed in the eyes) with a
chemical that might be toxic, you
should immediately call the NationalPoisonControlCenter for
advice at:
1-800-332-3073

Introduction
SafetyQuiz
Refertotheillustrationonthefollowingpagewhenansweringtheques
tions.
1. List three unsafe activities in the illustration and explain why
each is unsafe.
2. List three correct procedures depicted in the illustration.
3. What should Tarik do after the accident?
4. What should Lindsey have done to avoid an accident?
5. Compare Ming and David’s laboratory techniques. Who is
following the rules?
6. What are three things shown in the laboratory that should not
be there?

7. Compare Joe and Tyler’s laboratory techniques. Who is
working the correct way?
8. What will happen to Ray and Chris when the instructor
catches them?
9. List three items in the illustration that are there for the safety
of the students.
10. What is Consuela doing wrong?
Introduction
Introduction
Science lab Safety reinforcement Agreement

Any type of science experimentation involves potential hazards
and unforeseen risks may exist. The need
to prevent injuries and accidents cannot be over-emphasized!
Use of this lab manual and any LabPaq are expressly
conditioned upon the student agreeing to follow all
safety precautions and accept full responsibility for his or her
own actions. Study the safety section of the
manual until you can honestly state the following:
_ Before beginning an experiment, I will first read all directions
and then assemble and organize all required
equipment and supplies.
_ I will select a work area that is inaccessible to children and
pets while experiments are in progress. I will
not leave experiments unattended and I will not leave my work
area while chemical equipment is set up
unless the room will be locked.
_ To avoid the potential for accidents, I will clear my home-lab
workspace of all non-laboratory items
before setting up the equipment and supplies for my lab
experiments.

_ I will never attempt an experiment until I fully understand it.
If in doubt about any part of an experiment,
I will first speak with my instructor before proceeding.
_ I will wear safety goggles when working with chemicals or
items that get into my eyes.
_ I know that except for water, most solvents such as toluene,
alcohols, acetone, ethers, ethyl acetate, etc.
are highly flammable and should never be used near an open
flame.
_ I know that the heat created when water is added to
concentrated acids is sufficient to cause spattering.
When preparing dilute acid solutions, I will always add the acid
to the water (rather than the water to the
acid) while slowly stirring the mixture.
_ I know it is wise to wear rubber gloves and goggles when
handling acids and other dangerous chemicals,
that acid spills should be neutralized with sodium bicarbonate
(baking soda), and that acid spilled on the
skin or clothes should be washed off immediately with a lot of
cold water.

_ I know that many chemicals produce toxic fumes and that
cautious procedures should be used when
smelling any chemical. When I wish to smell a chemical I will
never hold it directly under my nose but
instead will use my hand to waft vapors toward my nose.
_ I will always handle glassware with respect and promptly
replace any defective glassware because even a
small crack can cause glass to break, especially when heated.
To avoid cuts and injuries, I will immediately
dispose of any broken glassware.
_ I will avoid burns by testing glass and metal objects for heat
before handling. I know that the preferred
first aid for burns is to immediately hold the burned area under
cold water for several minutes.
_ I know that serious accidents can occur if the wrong chemical
is used in an experiment. I will always
carefully read the label before removing any chemical from its
container.
_ I will avoid the possibility of contamination and accidents by
never returning an unused chemical to its
original container. To avoid waste, I will try to pour out only

the approximate amount of chemicals
required.
_ I know to immediately flush any chemical that spills on the
skin with cold water and then consult
a doctor if required.
Introduction
_ To protect myself from potential hazards I will wear long
pants, a long-sleeved shirt, and
enclosed shoes and I will tie up any loose hair, clothing, or
other materials when performing
chemical experiments.
_ I will never eat, drink, or smoke while performing
experiments.
_ After completing all experiments, I will clean up my work
area, wash my hands, and store the
lab equipment in a safe place that is inaccessible to children and

pets.
_ I will always conscientiously work in a reasonable and
prudent manner so as to optimize my
safety and the safety of others whenever and wherever I am
involved with any type of science
equipment or experimentation.
It is impossible to control students’ use of this lab manual and
related LabPaqs or students’ work
environments, the author(s) of this lab manual, the instructors
and institutions that adopt it, and
Hands-On Labs, Inc. the publisher of the manual and producer
of LabPaqs authorize the use of
these educational products only on the express condition that
the purchasers and users accept
full and complete responsibility for all and any liability related
to their use of same. Please review
this document several times until you are certain you understand
it and will fully abide by its
terms; then sign and date the agreement were indicated below.
I am a responsible adult who has read, understands, and agrees
to fully abide by all safety
precautions prescribed in this manual for laboratory work and

for the use of a LabPaq. Accordingly,
I recognize the inherent hazards potentially associated with
science experimentation; I will
always experiment in a safe and prudent manner; and I
unconditionally accept full and complete
responsibility for any and all liability related to my purchase
and/or use of a science LabPaq or any
other science products or materials provided by Hands-On Labs,
Inc. (HOL).
____________________________________________________
____________
Student’s Name (print) and Signature Date
Introduction
MSDS: Material Safety Data Sheets
A Material Safety Data Sheet (MSDS) is designed to provide
chemical, physical, health, and safety
information on chemical reagents and supplies. An important

skill in the safe use of chemicals is
being able to read an MSDS. It provides information about how
to handle store, transport, use,
and dispose of chemicals in a safe manner.
MSDS also provide workers and emergency personnel with the
proper procedures for handling
and working with chemical substances. While there is no
standard format for an MSDS, they
all provide basic information about physical data (melting point,
boiling point, flash point, etc.),
toxicity, health effects, first aid procedures, chemical reactivity,
safe storage, safe disposal,
protective equipment required, and spill cleanup procedures. An
MSDS is required to be readily
available at any business where any type of chemical is used.
Even day-care centers and grocery
stores need MSDS for their cleaning supplies.
It is important to know how to read and understand the MSDS.
They are normally designed and
written in the following sections:
Section1:ProductIdentification (Chemical Name and Trade
Names)

Section2:HazardousIngredients(Components and Percentages)
Section3:PhysicalData(Boiling point, density, solubility in
water, appearance, color, etc.)
Section4:FireandExplosionData(Flash point, extinguisher
media, special fire fighting procedures,
and unusual fire and explosion hazards)
Section5:HealthHazardData (Exposure limits, effects of
overexposure, emergency and first aid
procedures)
Section6:ReactivityData(Stability, conditions to avoid,
incompatible materials, etc.)
Section7:SpillorLeakProcedures (Steps to take to control and
clean up spills and leaks, and
waste disposal methods)
Section8:ControlMeasures (Respiratory protection, ventilation,
protection for eyes or skin, or
other needed protective equipment)

Section9:SpecialPrecautions(How to handle and store, steps to
take in a spill, disposal methods,
and other precautions)
Summary: The MSDS is a tool that is available to employers
and workers for making decisions
about chemicals. The least hazardous chemical should be
selected for use whenever possible, and
procedures for storing, using, and disposing of chemicals should
be written and communicated
to workers.
View MSDS information at www.hazard.com/msds/index.php.
You can also find a link to MSDS
information at www.LabPaq.com. If there is ever a problem or
question about the proper handling
of any chemical, seek information from one of these sources.
Introduction
LabPaq by

Hands-On Labs
experiments
observing Bacteria and
Blood
CynthiaAlonzo,M.S.
Version 42-0249-00-01
Review the safety materials and wear goggles when
working with chemicals. Read the entire exercise
before you begin. Take time to organize the materials
you will need and set aside a safe work space in
which to complete the exercise.
Experiment Summary:
Students will learn how to use a microscope to
observe prepared slides of three major types of
bacteria, the protists Paramecium and Amoeba,
yeast, and the fungi Penicillium. Students will
prepare slides to observe bacterial cultures obtained
from yogurt. They will also prepare and study blood

smears to identify platelets and red and white blood
cells.
ExpErimEnt
© Hands-On Labs, Inc. www.HOLscience.com 49
objectives
● Gain functional knowledge of microscope operations through
practical applications of a
microscope in the observation of bacteria and blood
● Identify and observe various bacterial shapes and
arrangements in a yogurt culture
● Identify and observe red and white blood cells in a blood
smear
Experiment Observing Bacteria and Blood

materials
MATEriAlS QTY iTEM DEScriPTioN
Student provides 1 Plain active culture yogurt
1 Collection container
1 Microscope with 100x oil immersion lens
1 Immersion oil
1 Toothpick
1 Bandage
1 Distilled water
LabPaq provides 1 Gloves, Disposable
1 Lens-paper-pack-50-sheets
1 Slide - Cover Glass - Cover Slip Cube
1 Lancet
1 Form, Lancet, Sterile – Directions for Use
1 Alcohol Prep Pad
4 Pipet, Long Thin Stem
1 Slide - Amoeba proteus
1 Slide - Anabaena, w.m.
1 Slide - Ascaris eggs, w.m.
1 Slide - Bacteria bacillus form

1 Slide - Bacteria coccus form
1 Slide - Bacteria spirillum
1 Slide - Letter e Focusing Slide
1 Slide - Paramecium conjugation
1 Slide - Penicillium w/conidia
1 Slide - Yeast, w.m.
1 Slide - Yogurt bacteria
1 Slide-Box-MBK with Blank-Slides
1 Mask, Face with Earloops
Note: The packaging and/or materials in this LabPaq may differ
slightly from that which is listed
above. For an exact listing of materials, refer to the Contents
List form included in the LabPaq.
Discussion and review
Since their invention in the late 1500s, light microscopes have
enhanced our knowledge of basic
microbiology, biomedical research, medical diagnostics, and

materials science. Light microscopes
can magnify objects up to 1500 times, revealing a world of
details unknown to the naked eye.
Light-microscopy technology has evolved far beyond the first
microscopes of Robert Hooke and
Antoni van Leeuwenhoek. Special techniques and optics have
been developed to reveal the
structures and biochemistry of living cells. Microscopes have
even entered the digital age, using
fluorescent technology and digital cameras.
A light microscope works similar to a refracting telescope with
some minor differences. A telescope
must gather large amounts of light from a dim, distant object.
Therefore, the telescope needs a
large objectivelens to gather as much light as possible and bring
it to a bright focus. Because
the objective lens is large, it brings the image of the object at a
distance to a focus, which is why
telescopes are much longer than microscopes. Then the
telescope eyepiece magnifies the image
as it brings it to your eye.
In contrast to a telescope, a microscope must gather light from a
tiny area of a thin, well-illuminated

specimen that is nearby. Hence, the microscope does not need a
large objective lens. Instead, the
microscope’s objective lens is small and spherical, which means
it has a much shorter focal length
on either side. The lens brings the image of the object into
focus at a short distance within the
microscope’s tube. Then a second lens, called an ocular lens or
eyepiece, magnifies the image as
it brings it to your eye.
The other major difference between a telescope and a
microscope is a microscope has a light
source and a condenser. The condenser is a lens system that
focuses the light from a source onto
a tiny, bright spot of the specimen, which is the same area the
objective lens examines.
Also, unlike a telescope, which has a fixed objective lens and
interchangeable eyepieces,
microscopes typically have interchangeable objective lenses and
fixed eyepieces. By changing
the objective lenses – moving from relatively flat, low-
magnification objectives to rounder, high-
magnification objectives – a microscope can bring increasingly
smaller areas into view. Light

gathering is not the primary task of a microscope objective lens,
as it is with that of a telescope.
The Parts of a light Microscope
A light microscope has the following basic systems:
Specimen control: used to hold and manipulate the specimen.
● Stage: where the specimen rests.
● clips: holds the specimen on the stage. When looking at a
magnified image, even moving the
specimen slightly can move parts of the image out of view.
llumination: used to shed light on the specimen. The simplest
illumination system is a mirror that
reflects room light up through the specimen.
● lamp: produces light. Typically, lamps are tungsten-filament
light bulbs. For specialized
applications, mercury or xenon lamps may be used to produce
ultraviolet light. Some
microscopes use lasers to scan the specimen.

● condenser: a lens system that aligns and focuses the light
from the lamp onto the specimen.
● Diaphragm or disc apertures: placed in the light path to alter
the amount of light reaching the
condenser. Varying the amount of light alters the image
contrast.
lenses: used to form the image.
● Objectivelens:gathers light from the specimen.
● Eyepiece: transmits and magnifies the image from the
objective lens to your eye.
● Nosepiece: a rotating mount that holds many objective
lenses.
● Tube: holds the eyepiece at the proper distance from the

objective lens and blocks out stray
light.
Focus: used to position the objective lens at the proper distance
from the specimen.
● coarse-focus knob: brings the object into the focal plane of
the objective lens.
● Fine-focus knob: makes fine adjustments to focus the image.
Support and alignment
● Arm: a curved portion that holds all of the optical parts at a
fixed distance and aligns them.
● Base: supports the weight of all of the microscope parts.
● Tube: connects to the arm of the microscope by way of a rack
and pinion gear, which allows
for focusing the image when changing lenses or observers and
moving the lenses away from
the stage when changing specimens.
Some of the parts mentioned previously vary among

microscopes. Microscopes come in two
basic configurations: upright and inverted. The microscope
shown in the Figure 1 is an upright
microscope, which has the illumination system below the stage
and the lens system above the
stage. An inverted microscope has the illumination system
above the stage and the lens system
below the stage. Inverted microscopes are better for looking
through thick specimens, such as
dishes of cultured cells, because the lenses can get closer to the
bottom of the dish where the
cells grow.
Figure 1: Upright microscope.
Light microscopes can reveal the structures of living cells and
tissues as well as of non-living
samples such as rocks and semiconductors. Microscopes can be
simple or complex in design,

and some can do more than one type of microscopy, each of
which reveals slightly different
information. The light microscope has greatly advanced our
biomedical knowledge and continues
to be a powerful tool for scientists.
Microscope Terms
● Depth of field: The vertical distance from above to below the
focal plane that yields an
acceptable image.
● Fieldofview:The area of the specimen that can be seen
through the microscope with a given
objective lens.
● Focal length: The distance required for a lens to bring the
light to a focus, (usually measured
in millimeters).
● Focalpoint/focus:The point at which the light from a lens
comes together.
● Magnification:The product of the magnifying powers of the
objective and eyepiece lenses.

For example, a 15x eyepiece and a 40x objective lens will give
you 600 power magnification
(15x x 40x = 600x).
● Numerical aperture: The measure of the lens’ light-collecting
ability.
● Resolution:The closest two objects can be before they are no
longer detected as separate
objects (usually measured in nanometers).
● ImageQuality: The quality of the microscope image is
assessed as follows:
● Brightness: How light or dark is the image? Brightness is
related to the illumination system.
The brightness can be changed by changing the wattage of the
lamp and by adjusting the
condenser diaphragm aperture. Brightness is also related to the

numerical aperture of the
objective lens; the larger the numerical aperture, the brighter
the image.
Figure 2: Pollen grain under proper brightness (left) and poor
brightness (right).
● Focus: Is the image blurry or well-defined? Focus is related
to focal length and can be controlled
with the focus knobs. The thickness of the cover glass on the
specimen slide can also affect
the ability to focus the image if it is too thick for the objective
lens. The correct thickness is
usually written on the side of the objective lens.
Figure 3: Pollen grain in focus (left) and out of focus (right).
● Resolution:How close can two points in the image be before
they are no longer seen as two
separate points? Resolution is related to the numerical aperture
of the objective lens – the
higher the numerical aperture, the better the resolution; and the
wavelength of light passing
through the lens – the shorter the wavelength, the better the
resolution.

Figure 4: Pollen grain with proper resolution (left) and poor
resolution (right).
● contrast: What is the difference in lighting between adjacent
areas of the specimen? Contrast
is related to the illumination system and can be adjusted by
changing the intensity of the
light and the diaphragm/pinhole aperture. Chemical stains
applied to the specimen can also
enhance contrast.
Figure 5: Pollen grain with proper contrast (left) and poor
contrast (right).
When specimens are observed by transmitted light, light must
pass through the specimen in order
to form an image. The thicker the specimen, the less light that
passes through, which creates a

darker image. Therefore, the specimens must be thin (0.1 to 0.5
mm). Many organic specimens
must be cut into thin sections before observation. Specimens of
rock or semiconductors are
too thick to be sectioned and observed by transmitted light, so
they are observed by the light
reflected from their surfaces.
Figure 6: Glial cell cultured from a rat brain.
Types of Microscopy
A major problem in observing specimens under a microscope is
that their images do not have
much contrast. This is especially true of living things, although
natural pigments, such as the
green in leaves, can provide good contrast. One way to improve
contrast is to treat the specimen
with colored pigments or dyes that bind to specific structures
within the specimen.
Different types of microscopy have been developed to improve
the contrast in specimens. The
specializations are mainly in the illumination systems and the
types of light passed through the

specimen. Brightfield is the basic microscope configuration, and
the images to this point are from
brightfield microscopes. This technique provides very little
contrast, and much of the contrast is
provided by staining the specimens. A darkfield microscope
uses a special condenser to block out
most of the bright light and illuminate the specimen with
oblique light, much like the moon blocks
the light from the sun in a solar eclipse. This optical setup
provides a totally dark background and
enhances the contrast of the image to bring out fine details of
bright areas at boundaries within
the specimen.
Following are various types of light microscopy techniques.
These techniques achieve different
results by using different optical components. The basic idea
involves splitting the light beam

into two pathways that illuminate the specimen. Light waves
that pass through dense structures
within the specimen slow down compared to those that pass
through less dense structures. As
all of the light waves are collected and transmitted to the
eyepiece, they are recombined, so they
interfere with each other. The interference patterns provide
contrast. They may show dark areas
(more dense) on a light background (less dense), or create a
type of false three-dimensional (3-D)
image.
● Phase-contrast: A phase-contrast microscope is best for
looking at living specimens, such as
cultured cells. The annular rings in the objective lens and the
condenser separate the light
paths. Light passing through the central part of the light path is
then recombined with light
traveling around the periphery of the specimen. Interference
produced by these two paths
produces images in which dense structures appear darker than
the background.
Figure 7: Phase contrast.

● Differential InterferenceContrast (DIC):DIC uses polarizing
filters and prisms to separate
and recombine the light paths, giving a 3-D appearance to the
specimen. DIC is also called
Nomarski after its inventor.
● HoffmanModulationContrast:Hoffman modulation contrast is
similar to DIC; however, it
uses plates with small slits in both the axis and the off-axis of
the light path to produce two
sets of light waves passing through the specimen. Again, a 3-D
image is formed.
● Polarization:The polarized-light microscope uses two
polarizers, one on either side of the
specimen, positioned perpendicular to each other so that only
light that passes through the
specimen reaches the eyepiece. Light is polarized in one plane
as it passes through the first
filter and reaches the specimen. Regularly spaced, patterned, or
crystalline portions of the

specimen rotate the light that passes through. Some of this
rotated light passes through
the second polarizing filter, so these regularly spaced areas
show up bright against a black
background.
● Fluorescence: This type of microscope uses high-energy,
short-wavelength light (usually
ultraviolet) to excite electrons within certain molecules inside a
specimen, causing those
electrons to shift to higher orbits. When they fall back to their
original energy levels, they
emit lower-energy, longer-wavelength light (usually in the
visible spectrum), which forms the
image.
CareandHandlingoftheMicroscope
● When moving a microscope, always use two hands. Place one
hand around the arm, lift the
scope, and put your other hand under the base of the scope for
support. Learning to carry

the scope in this way will force you to carry it carefully and
ensure you do not knock it against
anything while moving it.
● When putting the scope down, do so gently. If you bang your
scope down on the table,
eventually lenses and other parts will jar loose. The microscope
seems like a simple instrument,
but each eyepiece and objective is made up of a number of
lenses put together in a specific
way to create wonderful magnification. If you bang the scope
around, you are shaking upward
of 15 to 20 lenses.
● When handling the scope, always have clean hands. It would
be a shame to damage the scope
with too much peanut butter!
Storing the Microscope
● The best place to store the scope is on a sturdy desk, table, or
shelf where the scope will not
be disturbed. Make sure to keep the scope protected with a
plastic or vinyl cover when it is
not in use. Dust is an enemy to the lenses, so always cover the

scope.
● If you are unable to find a safe place where you can leave the
scope out, store it in its original
fitted, foam case packaging.
cleaning the Microscope
● The first step in keeping the microscope clean is to keep it
from getting dirty. Always keep the
microscope covered with the dust cover when it is not in use.
● The eyepiece will need cleaning from time to time. Due to its
position on the scope, it will
have a tendency to collect dust and oil from your eyelashes. The
eyepiece lens should be
cleaned with a high quality lens paper, available from a camera
shop or an eyeglass center.
Brush any visible dust from the lens and then wipe the lens.
Apply a bit of lens solution to the
lens paper to aid in cleaning. Use a cotton swab in place of lens
paper, but do not use facial
tissue to clean the lenses.
● Occasionally, the objective lenses will need cleaning. Use a

fresh area of lens paper for each
lens to avoid transferring dust from one lens to another.
● Clean the lenses in the glass condenser under the stage.
● Clean the glass lens over the light or the mirror, so an
optimal amount of light can shine
through. Follow up by wiping down the whole scope with a soft,
clean, cotton towel.
Using the Microscope
● Take the microscope body from the case. Put the eyepiece in
the opening in the tube at the
top of the microscope. Remove the objective lenses from their
individual containers and screw
them into the revolving nosepiece, placing each lens in its
respective color coded position.

● Adjust the tension on the focusing control knobs to suit your
touch or to compensate for
normal wear over time. To increase tension, hold the right-hand
knob firmly and turn the
opposite knob clockwise; turning the knob counterclockwise
loosens the tension.
● Unplug the rotating mirror bracket from the base of the
microscope, insert the mirror
(packaged separately with the microscope) into the bracket so
that it swivels freely, and plug
the bracket back into the base of the microscope.
● Tilt the arm of the microscope back until it is at a position
where you can comfortably look
into the microscope eyepiece.
● Place a slide under the clips on the stage with the area you
wish to view positioned between
the lens selected and the hole in the stage.
● Turn the nosepiece to select the longest lens (usually the
highest power lens). Lower the
barrel of the microscope with the coarse-focus knob until it
almost touches the slide. If the

barrel will not go that far, unscrew the focus stop-screw under
the arm of the microscope until
the lens can almost touch the slide. When the lens is in position,
lightly tighten the screw and
lock it in place with the knurled nut.
● Place a light source in front of the microscope; use the small
lever on the sub-stage condenser
to fully open the diaphragm; and adjust the mirror so the light is
brightest when seen through
the microscope.
● Rotate the nosepiece to select the lowest power lens. Lower
the barrel with the coarse-focus
knob until the tip of the lens is near the slide. Now raise the
barrel slowly with the coarse-
focus knob until you see an image from the slide. Finish the
focus with the fine-focus knob.
● With your thumb and forefinger on each end of the slide,
move it slowly on the stage until the
object you wish to study is centered in your field of view.
● Rotate the nosepiece of the microscope to select the objective
lens that will give you the

higher magnification you need.
● Once one lens is focused properly, any other objective lens
on the nosepiece when rotated
into position will be roughly in focus and require only fine
focus to bring the image into correct
focus.
● Move the lever for the diaphragm through its full range to
select the amount of light that gives
you the best contrast. Many details will be visible with good
contrast which would otherwise
be lost with too much or too little light.
Using the Electric illuminator
With your fingers, grasp the illuminating mirror behind its
bracket and pull to unplug the bracket
and mirror from the base of the microscope. Insert the metal

plug tip of the electric illuminator
into the hole from which you unplugged the mirror bracket.
Rotate the fixture so that the glass
opening over the bulb points up toward the light condenser
under the stage. Plug the electric
cord into a 115-volt outlet and turn on the switch in the cord.
UsingtheOilImmersionLens(purchasedseparately)
Install the oil immersion 100x objective lens in place of any of
the other objective lenses. The 4x
lens is a good choice. First, focus the microscope and center the
slide using a lower magnification
objective. Apply a drop of oil on the specimen slide and turn the
revolving nosepiece to bring the
100x objective into position. If the barrel is too low to allow the
100x lens to move into position,
raise it very slightly with the coarse focus, position the lens,
and then lower the barrel until the
tip of the 100x lens touches the oil. The tip of the lens is able to
move a short distance into the
lens against a spring in order to keep from putting too much
pressure on the slide. With the lens
tip touching the oil, focus with the fine-focus knob. The
working distance of the lens is very short,

so do not use the coarse-focus knob other than to position the
lens. After using the oil immersion
lens, wipe off the oil carefully with alcohol.
Exercise1:ViewingPreparedSlides
procedure
PartI:ViewingPreparedSlides
1. Set up your microscope. Refer to the Discussion and Review
section for more information.
2. Clean the ocular lenses and objectives with lens paper prior
to use.
3. Place the prepared e focusing slide, cover slip up, on the
stage within the spring loaded lever.
4. Turn the rotating nosepiece until the 10x objective is above
the ring of light coming through

the slide.
5. Move the slide using the X and Y stage travel knobs until the
specimen is within the field of
view.
6. Adjust the focus by looking into the eyepiece and focusing
the specimen with the coarse then
fine focus knobs.
7. Bring the condenser up to the bottom of the slide and then
slightly back for maximum light.
8. Adjust the iris diaphragm until there is sufficient light
passing through the specimen. This
will take practice. Begin with the diaphragm closed and slowly
open it while observing the
specimen. Choose the level at which there is enough light to
allow good resolution, but not so
much light that there is a glare or whitening of the field of
view.
9. Repeat the previous steps with six different prepared slides
with 10x and 40x objectives. Refer
to Figure 8 for image comparisons.

Figure 8: Comparisons of slides with 10x and 40x objective
lenses.
Fungi – 10x lens Fungi – 40x lens
Yeast – 10x lens Yeast – 40x lens
Paramecium – 10x lens Paramecium – 40x lens
Part ii: Using an immersion oil lens
The most important objective used in microbiology is the oil
immersion lens, 100x. Many bacteria
cannot be visualized clearly without the use of oil immersion.
When using an oil immersion lens,

oil is placed between the objective and the slide to prevent the
loss of light due to the bending of
light rays as they pass through air. This enhances the
resolvingpower of the microscope.
1. After focusing with a high, dry objective, turn the 40x
objective away from the specimen.
2. Place a drop of oil on the slide.
3. Rotate the oil immersion objective, 100x, into the oil, then
past the oil and back. This ensures
there are no air bubbles between the objective and the oil.
4. Use only the fine focus to bring the object into focus.
5. Practice viewing at least six prepared slides at 10x, 40x, and
100x with oil. Refer to Figure 9
for image comparisons.
a. When replacing slides on the stage, start with the 10x or the
40x before going to oil. Do
not let oil get on the 10x and 40x objectives.

b. Always rotate the oil immersion objective away before
removing a slide.
c. Never use the coarse focus with the oil objective in place.
The slide could break and the
objective could get damaged.
6. Clean the oil off the oil objective with lens paper. Then clean
all the objectives with clean lens
paper.
Figure 9: Comparisons of slides with the 10x, 40x, and 100x
(oil immersion) lenses
Yeast – 10x lens Yeast – 40x lens
Yeast - 100x oil immersion lens
Fungi – 10x lens Fungi – 40x lens

Fungi - 100x oil immersion lens
Questions
A. Identify the following parts of the microscope and describe
the function of each.
B. Define the following microscopy terms:
● Focus:
● Resolution:
● Contrast:
C. What is the purpose of immersion oil? Why does it work?

Exercise 2: observing Bacteria cultures in yogurt
Bacteria occur in a variety of different shapes. By far, the most
numerous are spheres, rods,
commas, and spirals. Spherical bacteria, called cocci, and rod
shaped bacteria, called bacillus, are
the most common shapes.
Figure 10: Bacteria shapes.
In addition to shape, the way individual bacteria are arranged is
an identifying feature. For
example, bacteria can occur in pairs (diplo), strands (strepto),
or clusters (staphylo). A common
inhabitant of yogurt is a paired, round bacteria – diplococcus.
Figure 11: Bacteria arrangements.

procedure
1. Locate a small, sealable container made of glass or plastic.
Clean the container thoroughly
with soap and then rinse the container several times to remove
all the soap.
2. Place a teaspoon of yogurt in the container.
3. Cover the container and place it in a dark, relatively warm
area. Leave the container
undisturbed for 12–24 hours.
4. Use a toothpick to take a sample of yogurt from the container
and place the sample on a clean
slide. If the sample on the slide seems too thick, dilute it with a
drop of water.
5. Place a cover slip on top of the sample.
6. Observe the bacteria under the microscope at 10x, 40x, and
100x oil immersion. The diaphragm
setting should be very low, because the fresh bacteria will
appear nearly transparent.

7. Next, view the prepared stained yogurt slide from the kit.
Compare your observations of the
fresh, live slide to the prepared, stained slide.
8. Clean the collection vials and slides thoroughly after use.
Questions
A. Describe your observations of the fresh yogurt slide.
B. Were there observable differences between your fresh yogurt
slide and the prepared yogurt
slide? If so, explain.
C. Describe the four main bacterial shapes.
D. What are the common arrangements of bacteria?
E. Were you able to identify specific bacterial morphologies on
either yogurt slide? If so, which
types?

Exercise 3: Preparing and observing a Blood Slide
procedure
Part i: Preparing a Blood Slide
WArNiNg: Blood can carry diseases that can be transferred
from person to person. Avoid contact
with another person’s blood. When necessary to contact blood,
wear rubber gloves.
1. Thoroughly wash your hands with soap and warm water.
2. Clean a finger tip with the alcohol prep pad and allow to dry.
3. Quickly and lightly poke the inside of your sterilized finger
with the lancet.
4. Squeeze your finger to place a drop of blood on a clean slide
in accordance with the following
directions.
a. Drop the blood toward one end of a slide as shown in Figure

12.
b. Tilt the cover slip toward the drop. Then slowly move the
slip toward the drop until it
contacts the blood and grabs the drop.
c. Without changing the tilt of the cover slip, move the slip
back over the slide, drawing the
blood across the slide.
d. Lay the cover slip flat across the blood smear.
Figure 12: Blood smear preparation.
5. Place a bandage on your finger to prevent infection.
Part ii: observing Blood
Human blood appears to be a red liquid to the naked eye, but

under a microscope it contains four
distinct elements:
● plasma
● red blood cells
● white blood cells
● platelets
The plasma is the liquid part of blood and is actually straw-
yellow in color. The red blood cells give
blood its red color. White blood cells are interspersed in the sea
of red blood cells and help fight
infection. The platelets are fragments of red blood cells and
function in clotting. While red blood
cells should be visible on the slide, white blood cells and
platelets may be harder to find.
1. Place the blood slide on the microscope stage and bring it
into focus on low power. Adjust
the lighting and then switch to a 40x magnification. To view
individual cells, use 100x oil
immersion.

You should see hundreds of tiny red blood cells. There are
billions circulating throughout your
blood stream. Red blood cells contain no nucleus, which means
they can’t divide. Red blood cells
are constantly produced by the bone marrow and the spleen.
You should also be able to find a few white blood cells. They
are slightly larger than red blood
cells and have a nucleus. Some, macrophages, often resemble an
amoeba and can contort their
body in any way they like to engulf foreign objects. Others are
spherical. White blood cells fight
infection by consuming foreign bodies or injecting them with
enzymes that induce cell death or
apoptosis. Platelets are fragments of red blood cells and are
very small.
Figure 13: Blood smear slides at 10x, 40x, and 100x (oil
immersion) lenses.
10x lens 40x lens
100x lens

Questions
A. Describe the cells you were able to see in the blood smear.
B. Are the cells you observed in your blood smear different than
the bacterial cells you have
observed? Why or why not?
observing Bacteria and Blood
CynthiaAlonzo,M.S.
Version 42-0249-00-01
lab report Assistant
This document is not meant to be a substitute for a formal

laboratory report. The Lab Report
Assistant is simply a summary of the experiment’s questions,
diagrams if needed, and data tables
that should be addressed in a formal lab report. The intent is to
facilitate students’ writing of lab
reports by providing this information in an editable file which
can be sent to an instructor.
Exercise1:ViewingPreparedSlides
Questions
A. Identify the following parts of the microscope and describe
the function of each.
Define the following microscopy terms:
● Focus: Is the image blurry or well-defined?
● Resolution:

● Contrast:
B. What is the purpose of immersion oil? Why does it work?
Exercise 2: observing Bacteria cultures in yogurt
Questions
A. Describe your observations of the fresh yogurt slide.
B. Were there observable differences between your fresh yogurt
slide and the prepared yogurt
slide? If so, explain.
C. Describe the four main bacterial shapes.
Cocci –
Bacillus –
Spirillum –

Vibrio –
D. What are the common arrangements of bacteria?
Diplo –
Strepto –
Staphylo -
E. Were you able to identify specific bacterial morphologies on
either yogurt slide? If so, which
types?
Exercise 3: Preparing and observing a Blood Slide
Questions
A. Describe the cells you were able to see in the blood smear.
B. Are the cells you observed in your blood smear different than
the bacterial cells you have
observed? Why or why not?

Bacterial Morphology
CynthiaAlonzo,M.S.
Version 42-0240-00-01
Review the safety materials and wear goggles when
working with chemicals. Read the entire exercise
before you begin. Take time to organize the materials
you will need and set aside a safe work space in
which to complete the exercise.
Experiment Summary:
Students will observe various bacterial morphologies
using prepared slides. They will prepare live culture
smears of Saccharomyces cerevisiae and cheek cells,
and view these specimens under a microscope using
direct and indirect staining techniques. Students will
also learn how to prepare disinfectants and use them
to decontaminate working surfaces.
ExpErimEnt

© Hands-On Labs, Inc. www.HOLscience.com 73
objectives
● Observe bacterial morphologies by preparing wet-mount
slides
● Learn and employ direct and indirect staining techniques
Experiment Bacterial Morphology
materials
MATEriAlS QTY iTEM DEScriPTioN
Student provides 1 10%-bleach solution
1 Microscope
1 Immersion Oil
3 Toothpicks

1 Warm water
1 Paper towels
1 Clothespin, tweezers, or test tube holder
LabPaq provides 1 Gloves, disposable (1 pair)
1 Goggles, safety
1 Apron, plastic
1 Slide – Cover Glass – Cover Slip Cube (3)
1 Lens-paper-pack-50-sheets
1 Cup, Plastic, 9 oz Tall
1 Pencil, marking
1 Tray-Staining tray
2 Candle, tea size (flame source)
1 Congo Red Stain, 0.1% - 1 mL in Pipet
2 Baker’s Yeast Packet – Saccharomyces cerevisiae
2 Pipet, Long Thin Stem
1 Gram Stain

Laboratory Format for Lab Reports Title Page (Contains ti.docx

More Related Content

Similar to Laboratory Format for Lab Reports Title Page (Contains ti.docx (20)

More from DIPESH30 (20)

Recently uploaded (20)

Laboratory Format for Lab Reports Title Page (Contains ti.docx