![Glucose
•↑ CSF glucose conc. (hyperglycorrhachia) :
•Not clinically informative
•Provides only confirmation of hyperglycemia
•↓CSF [glucose] (hypoglycorrhachia):
1)Disorder in carrier-mediated transport e.g. TB meningitis,
2)Active metabolism of glucose by cells or organisms:
•e.g. acute purulent, amebic, & fungal meningitis
3)Increased metabolism by the CNS
4)Glucose is measured by the use of a Spectrophotometer
7/22/2019 36](https://image.slidesharecdn.com/laboratoryanalysisofcsf-190722131552/85/Laboratory-analysis-of-csf-36-320.jpg)
Laboratory analysis of csf
- 1. UNIVERSITY OF BUEA 7/22/2019 1 Faculty of Health Sciences Department of MLS Chemical Pathology: CPY 602, BIOCHEMICAL ANALYSISOF CEREBRO-SPINALFLUID Presented By Lamngwa Benard Nfor HS18P036
- 2. OUTLINE Introduction Formation of CSF/anatomy Circulation of CSF Functions of CSF Blood brain barrier Biochemical composition of CSF CSF analysis and pathophysiology Laboratory investigations Management of some CSF related infections 7/22/2019 2
- 3. OBJECTIVES After this presentation, the students should be able to: Describe the formation and location of cerebrospinal fluid (CSF) Describe the appearance and state the composition of normal CSF Understand the formation and functions of CSF Understand some pathology associated to CSF Understand biochemical investigations of CSF and interpretation of such results Have an idea of management of some related disease 7/22/2019 3
- 4. INTRODUCTION •The cerebrospinal fluid (CSF) is a dynamic, metabolically active fluid surrounding the brain and spinal cord and has many important functions. •It is very valuable as a diagnostic aid in the evaluation of inflammatory conditions, infections involving the brain, spinal cord, and subarachnoid haemorrhage. 7/22/2019 4
- 5. FORMATION OF CSF/ANATOMY •Both the brain and spinal cord are covered by three protective membranes referred to as the meninges. The outermost layer is called the dura mater and is composed of tough connective tissue. •The middle layer is the arachnoid named for it spider web like appearance. •The delicate innermost layer which is in direct contact with the brain and spinal cord is called the pia mater. An inflammation of the meninges is referred to as meningitis. 7/22/2019 5
- 6. FORMATION OF CSF ANATOMY CONT. •Between the arachnoid layer and the pia mater is a space called the subarachnoid space. It contains a clear, colorless fluid referred to as Cerebrospinal Fluid (CSF). •CSF is produced in the ventricles of the brain by a collection of rich vascular protrusions called the choroid plexus. •Excess CSF is continuously reabsorbed by arachnoid villi and returned to the venous system thus maintaining a consistent amount of fluid under an intracranial pressure between 50 - 180 mmHg. 7/22/2019 6
- 9. FORMATION OF CSF /ANATOMY CONT. •Generally, the total volume of CSF circulating throughout the adult Central Nervous System (Brain and Spinal Cord) is approximately 90 - 150 ml. •In newborns this volume is 10 -60 ml. 7/22/2019 9
- 10. CIRCULATION OF CSF •CSF flows from the lateral ventricles through the third and fourth ventricles and into the subarachnoid space. • From the fourth ventricle, the CSF either continues to the central canal of the spinal cord or leaves for the subarachnoid space by passing through the medial aperture (foramen of Magendie) and paired lateral apertures (foramina of Luschka) in the roof and lateral recesses of the fourth ventricle, respectively. 7/22/2019 10
- 11. CIRCULATION OF CSF •CSF flows from the lateral ventricles through the third and fourth ventricles and into the subarachnoid space. • From the fourth ventricle, the CSF either continues to the central canal of the spinal cord or leaves for the subarachnoid space by passing through the medial aperture (foramen of Magendie) and paired lateral apertures (foramina of Luschka) in the roof and lateral recesses of the fourth ventricle, respectively. 7/22/2019 11
- 12. FUNCTIONS OF CSF •Protection : CSF protects the brain from damage by buffering the brain. It acts as a cushion •Buoyancy: The actual mass of the human brain is about 1400 grams; however, the net weight of the brain suspended in the CSF is equivalent to a mass of 25 grams. which allows the brain to maintain its density without being impaired by its own weight. 7/22/2019 12
- 13. FUNCTIONS OF CSF cont. •Chemical stability/Homeostasis: CSF maintain the distribution of necessary substance and waste product between CNS and Blood stream •Prevention of brain ischemia: made by decreasing the amount of CSF in the limited space inside the skull. This decreases total pressure . transport of biomolecules to the brain •Clearance of catabolites (CO2, lactate) •Maintenance of constant intracranial pressure •Clearin of waste: Removes waste from the brain through the bllod for elimination via kidneys. 7/22/2019 13
- 14. BLOOD BRAIN BARRIER •. A protective barrier separates the brain from circulating blood and regulates the distribution of substances between the blood and the CSF. • If this protective barrier disrupts, it may result in a change in the normal level or type of constituents of CSF. •The blood–brain barrier restricts the passage of pathogens, the diffusion of solutes in the blood, and large or hydrophilic molecules into the cerebrospinal fluid (CSF), while allowing the diffusion of hydrophobic molecules (O2, CO2, hormones) and small polar molecules. •Cells of the barrier actively transport metabolic products such as glucose across the barrier using specific transport proteins.7/22/2019 14
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- 16. BIOCHEMICAL COMPOSITION OF CSF 7/22/2019 16
- 17. Collection of CSF •The patient is placed in a lateral position with the knees bent in full flexion up to the chest(fetal position) (sit and bend position also possible) •Introduction of 1% lidocain into the subcutaneous space •Insertion of a spinal needle into the subarachnoid space at the L3-L4 or L4-L5 intercostal spaces (inserted until the second “give”- subarachnoidal space). •Removal of the stylet of the needle in order to collect the fluid 7/22/2019 17
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- 20. Collection of CSF • Approximately 10-12 ml can be taken from an adult and less than 3 ml from a neonate. Care must be taken not to contaminate the sample with blood. •The fluid is collected into 3 tubes via free passive flow through the needle. Up to four tubes of CSF can be collected aseptically. •Each tube should contain 2-4 mL of fluid. The tubes should be labelled with a number indicating the order in which they were collected. Each tube goes to a different section of the laboratory for testing 7/22/2019 20
- 21. Collection of CSF • Tube #1 Chemistry: Contains debris from the puncture and occasionally blood in a "traumatic tap". Since it is the most likely to be contaminated with microbes, tissue fluid and blood cells which could yield misleading results. It is best used for chemistry and immunological determinations. • Tube #2 Microbiology. May contain some blood cell contaminants but is suitable for microbiological studies. • Tube #3 Haematology. Has the least cellular or debris contamination and therefore is used for cell counts, white cell differentials and the examination of abnormal cells e.g. tumor cells • Tube #4, Molecular & Reference Lab7/22/2019 21
- 22. CSF ANALYSIS AND PATHOPHYSIOLOLOGY • CSF is investigated to aid in the diagnosis of SAH (subarachnoid hemorrhage), Meningitis, Demyelinating disorders, CNS malignancy, Head injury with CSF leak and other less common tests, the majority of the investigations involve cytology and microbiology and chemistry. •Meningitis •Can be bacterial ( pyogenic or tuberculous), viral or fungal ( Cryptococcus). Apart from direct organism identification, biochemistry and cell counts can help distinguish between the types of meningitis. 7/22/2019 22
- 23. CSF ANALYSIS AND PATHOPHYSIOLOGY cont. • In classical pyogenic meningitis ( eg Strep Cryptococcal) CSF protein levels are high > 0.8 g/L, glucose is much lower than 60% of blood value, high counts of PMNL’ s ( polymorphonuclear lymphocytes) occur and the CSF is turbid. •TBM (tuberculous meningitis) can be differentiated by a mixed PMNL/lymphocyte high cell count and a low CSF chloride. •Viral meningitis often has a lymphocytosis with a normal glucose, normal protein levels and a clear CSF. •Fungal infections present with CSF lymphocytosis, a high protein count and normal glucose 7/22/2019 23
- 25. CSF ANALYSIS AND PATHOPHYSIOLOGY cont. • CSF GLUCOSE •Glucose enters the CSF from the plasma by 2 mechanisms, diffusion and active transport. Glucose is the primary source of energy for the brain. Brain is 5 % body mass but uses 20% glucose. •The level of CSF glucose is influenced by the concentration and duration of the plasma glucose level. •CSF glucose levels are roughly two-thirds of plasma glucose levels •CSF glucose can only be interpreted reliably with a blood sample taken within 15 min before or after an LP. 7/22/2019 25
- 26. CSF ANALYSIS AND PATHOPHSIOLOGY cont. CSF CHLORIDE The diagnosis of tuberculous meningitis is notoriously difficult to make. Chloride (less than 110mmol/1) is used to distinguish tuberculous from viral and bacterial meningitis. A CSF chloride of less than 100mmol/l is thought to be virtually diagnostic of TBM. CSF PROTEIN. Spinal fluid is an ultrafiltrate of plasma that lacks high molecular weight proteins such as beta lipoprotein, alpha-2 macroglobulin, IgM, etc. 7/22/2019 26
- 27. CSF ANALYSIS AND PATHOPHYSIOLOGY cont. Measurement of CSF protein is also used to distinguish septic from aseptic meningitis. Protein concentrations >1 g/L are often viewed as diagnostic for bacterial, fungal, or tuberculous meningitis, reflecting the presence of the pathologic organism Unfortunately, the increase of CSF protein is a sequela in many CNS disease processes, usually in association with increased permeability of the bloodbrain barrier, vasogenic brain edema, hypercellularity, and release of brain-specific proteins during cell death 7/22/2019 27
- 28. CSF ANALYSIS AND PATHOPHYSIOLOGY cont. SUBARACHNOID HAEMORRHAGE In subarachnoid haemorrhage red cells are a diagnostic feature in the CSF, so careful LP is essential. Red cells that have been in the CSF for longer than 4 hours cause a yellow staining of the CSF called xanthochromia and the presence of this in fresh CSF confirms bleeding into the subarachnoid space from a source other than contamination during the LP. Keep in mind that there are other causes of xanthochromia. Xanthochromia refers to a yellow, orange or pink colour7/22/2019 28
- 29. CSF ANALYSIS AND PATHOPHYSIOLOGY cont. HEAD INJURY • In patients with rhinorrhoea or otorrhoea, post head injury or spontaneously, it is important to ascertain whether CSF is present in the fluid, which would confirm a CSF leak. This is done by identifying beta- 2-transferrin in the fluid leak sample, using electrophoresis and immunofixation 7/22/2019 29
- 30. CSF ANALYSIS AND PATHOLOGY cont. • Protein index: • This assesses the amount of intrathecal protein synthesis that may occur in an inflammatory disease and assesses the permeability of the blood-brain barrier in relation to increased intrathecal synthesis (eg IgG). • Albumin is used as a reference protein. • A normal CSF Albumin/Serum Albumin ratio is less than 9. CSF IgG index = CSF IgG/Serum IgG ÷ CSF Albumin/Serum Albumin Usually the CSF IgG index will be 0.3-0.8. If > 0.8, this indicates increased intrathecal synthesis such as may be seen in multiple sclerosis.. 7/22/2019 30
- 31. LABORATORY INVESTIGATIONS • Physical examination •Normal CSF is: •Colorless •Clear •Free of clots •Free of blood •If CSF is cloudy (turbid) , perform microscopic examination: •Usually due to leucocytes •may be due to micro- organisms 7/22/2019 31
- 32. LABORATORY INVESTIGATIONS • Physical examination •Normal CSF is: •Colorless •Clear •Free of clots •Free of blood •If CSF is cloudy (turbid) , perform microscopic examination: •Usually due to leucocytes •may be due to micro- organisms 7/22/2019 32
- 33. Blood & Hemoglobin pigments in CSF Traumatic tap •bright red color • RBCS in decreasing number as the fluid is sampled Subarachnoid hemorrhage (SAH) •Xanthochromia (hemoglobin breakdown pigments) = RBCs lysis & metabolism previously occurred (at least 2 hr earlier) 7/22/2019 33
- 34. Examination of CSF (Biochemical analysis of CSF) •Tests of interest: •Glucose •Protein Total Specific: Albumin Immunoglobulin Others (e.g. myelin basic protein; MBP) Lactate 7/22/2019 34
- 35. Glucose • Glucose enters CSF via facilitative transporter (GLUT) • CSF glucose is ~ 2/3 that of plasma 50 - 80 mg/dl • A plasma sample must be obtained ~ 2-4 hr before CSF sample • In hypoglycemia, CSF glucose may be very low and In hyperglycemia glucose is raised. • Measure CSF Glucose immediately or preserve the specimen with and antiglycolytic 7/22/2019 35
- 36. Glucose •↑ CSF glucose conc. (hyperglycorrhachia) : •Not clinically informative •Provides only confirmation of hyperglycemia •↓CSF [glucose] (hypoglycorrhachia): 1)Disorder in carrier-mediated transport e.g. TB meningitis, 2)Active metabolism of glucose by cells or organisms: •e.g. acute purulent, amebic, & fungal meningitis 3)Increased metabolism by the CNS 4)Glucose is measured by the use of a Spectrophotometer 7/22/2019 36
- 37. Proteins •The Proteins mostly found in the CSF is albumin •Source of CSF proteins: • 80% from plasma by ultrafiltration • 20% from intrathecal synthesis 7/22/2019 37
- 38. proteins cont. •Increase CSF total protein Must be compared to the serum protein conc. Useful nonspecific indicator of pathological states: •Lysis of contaminant blood (traumatic tap) •↑ Permeability of the epithelial membrane due to: •Bacterial or fungal infection, cerebral hemorrhage •↑ production by CNS tissue in, e.g Multiple sclerosis (MS) •Obstruction e.g. in tumors and abscess 7/22/2019 38
- 39. Proteins cont. •Total protein is measured spectrophotometrically •The Pandys test is also used for the qualitative detection of total protein in CSF 7/22/2019 39
- 40. Albumin • Albumin is produced solely in the liver • Its presence in CSF must occur through BBB • Measured by the protein electrophoresis method 7/22/2019 40
- 41. Immunoglobulin • CSF IgG can arise: • from plasma cells within CSF • from the blood through BBB •Increase IgG conc. and normal Alb conc. of CSF suggests local production of IgG, e.g., • Multiple sclerosis (MS) • Subacute sclerosing panencephalitis (SSPE) • IgG can be measured by electrophoretic method • Also measured by immunoassays. 7/22/2019 41
- 42. Management of CSF related infections Meningitis Bacterial meningitis is a medical emergency Empirical selection of therapy is usually necessary, and treatment should be started at the first suspicion of bacterial meningitis. A single dose of benzylpenicillin can be given if the person is outside hospital, but cefotaxime is the preferred treatment in hospital Chloramphenicol is an option for those who have an allergy to both penicillin and cephalosporins. Treatment is given for 5 days for meningococcus, and 10 days for Haemophilus influenzae or pneumococcus. 7/22/2019 42
- 43. Management of CSF related infections Meningitis Amphotericin is active against all common fungi that cause systemic infection (Candida, Aspergillus, Mucor and Cryptococcus species). Cryptococcus is incriminated in cryptococcal meningitis. Tuberculosis is usually treated with a multidrug regimen because of the rapid development of resistance. Rifamycins (rifabutin, rifampicin), Isoniazid, Pyrazinamide and Ethambutol are first line regimen. Other drugs can be used as second-line treatments in multidrug- resistant tuberculosis. These include cycloserine, capreomycin, amikacin, ciprofloxacin, moxifloxacin, azithromycin, clarithromycin and p-aminosalicylic acid7/22/2019 43
- 44. Management of CSF related infections Acute brain injury Mannitol is occasionally used to reduce ischaemic cerebral damage, for example after neurosurgery or in acute traumatic brain injury. Fluid loss via the kidney should be replaced with intravenous crystalloid to avoid dehydration. Dexamethasone is often used to reduce oedema around malignant tumours in the brain and those compressing the spinal cord 7/22/2019 44
- 45. REFERENCES 1. Cerebrospinal fluid analysis in infectious diseases of the nervous system: when to ask, what to ask, what to expect, Luís dos Ramos Machado1,3, José Antonio Livramento2,3, Liliana Scaff Vianna4 2. Scott LA, Tintinalli JE, Brewer KL, Salgado CD. Lumbar punctures for 3. suspected meningitis in adults. Infect Dis Clin Pract 2008;16:298-302. 4. 2. Van de Beek D, de Gans J, Tunkel AR, Wijdicks EFM. Communityacquired bacterial meningitis. N Engl J Med 2006;354:44-53Dalia Mohamed Moemen 5. Cerebrospinal fluid, Assistant lecturer of Medical Microbiology and Immunology, Medical Microbiology and Immunology Department, Faculty of Medicine, Mansoura Universit 6. Clinical Utility of Biochemical Analysis of Cerebrospinal Fluid Mark A. Watson and Mitchell G. ScottRoutine c erebrospinal fluid ( CSF ) a nalysis 7. F. Deisenhammer,1 A. Bartos,2 R. Egg,1 N. E. Gilhus,3 G. Giovannoni,4 S. Rauer,5 Sellebjerg,6 H. Tumani 8. CSF ANALYSIS IN METABOLIC DISORDERS© CSF Neurotransmitter Request Form (.pdf)7/22/2019 45
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