Left to right, Normal CSF, mildly xanthochromicCSF, moderately xanthochromic CSF, redtingedturbid CSF caused by hemorrhage, and cloudyred-tinged fluid from a horse with bacterial meningitis.
Mono, lympho, neutro
Macro. Lumpho, neuto
Examination of cerebrospinal fluid presentation mode
Examination of CerebrospinalfluidDr.Pavulraj.S5246M.V.Sc., scholarDivision of pathologyIndian Veterinary Research InstituteIndia
Introduction• Cerebrospinal Fluid - clear, colorless transparent,tissue fluid present in the cerebral ventricles, spinalcanal and subarachnoid spaces.• Almost no blood cells, little protein and more salt
Formation of cerebrospinal fluid• CSF is largely formed by the choroid plexus of the lateralventricle and remainder in the third and fourth ventricles.• 30% of the CSF is also formed from the ependymal cellslining the ventricles and other brain capillaries.• The choroid plexus of the ventricles actively secretecerebrospinal fluid.• The choroid plexuses are highly vascular tufts covered byependyma.
Circulation of CSFSubarachnoid space of Brain and Spinal cordForamen of Megendie and foramen of LuschkaFourth ventricleCerebral aqueduct of SylviusThird ventricleForamen of Monro [Interventricular foramen]Lateral ventricle
Function of CSF• Mechanical cushion to brain• Source of nutrition to brain• Excretion of metabolic waste products• Intra-cerebral transport medium• Control of chemical environment• Auto-regulation of intracranial pressure
Indications• Diagnostic purpose– Infections: meningitis, encephalitis– Inflammatory conditions– Infiltrative conditions : Leukemia, lymphoma,carcinomatous - meningitis History of seizures or CNS diseases. Relief of abnormally high pressure and drainageof blood or exudate. As a prognostic tool for evaluation of CNSdiseases. To assess the response to treatment.
CollectionSite• Cisterna magna or Atlanto-occipital puncture - Horse,cat & dog.• Sub lumbar or Lumbosacralpuncture - cow, sheep &goat.• Only Lumbar puncture – PigInstruments• 12.5 cm long 14G needlewith a stylet – Large animal• 3 inch long 16 G spinalneedle – Small animal
Normal components of CSFNormal biochemical constituents of CSF: Lower in CSF than plasma Proteins Glucose Phosphorus Bicarbonate Potassium Sulfate Cholesterol Enzymes Higher in CSF than plasma Sodium Chloride CSF normally does not contain erythrocytes. Normal CSF consists of varying proportions of small lymphocytes and monocytes. Major protein in CSF is albumin. The major Ig in normal CSF is IgG, which normally originates from the serum. The normal CSF glucose level is about 60% to 80% of the blood glucose concentration.
Examination of CSFPhysical examinationChemical examinationCytological examinationBacteriological examination• Increased glucose level in the CSF-hyperglycorrhacia• Decreased glucose level in the CSF -hypoglycorrhacia• Increased number of white blood cells in CSF -pleocytosis
Macroscopic examination• Color– Clear and colorless as distilled water• Normal• Encephalitis and meningitis associated with viralinfections– Bright red• Puncture of blood vessels• Old hemorrhage (yellow supernatant)– Brown or dull red• Intra cranial hemorrhage– Yellow• Xanthochromic – bilirubin from disintegration ofRBC in subarachnoid space from old hemorrhage• Excess bilirubin in plasmaXanthochromia
• Turbidity– Due to presence of cells - >500/μl– Bacterial meningitis, hemorrhage• Coagulation– Normal CSF – not coagulate– Increased protein- fibrinogen– Acute suppurative meningitis• Specific gravity- 1.003-1.008• Reaction – Alkaline as like blood
Chemical examination• Protein - Normal range – 10-40mg/dl– Present in very small quantity – albumin– Globulin – pathological conditions• Total protein• Foam test• Sulfasalicylic acid test• Globulin• Nonne-Apelt test – 1ml ammonium sulfate +1ml CSF – Gray ring• Pandy’s test – 1ml phenol+1ml CSF – Turbidity• Increased level– Inflammation – meningitis, encephalitis– Neoplasia– Hemorrhage– Uremia– Tissue destruction
Cytological ExaminationTotal cell count Collection of CSF in plastic or silicon coatedglass tube is preferred. The total cell counts of the CSF must beestimated within 20 minutes of collection,since the cells degenerate rapidly. Storage can be done at 4–8 C (short term) orat −20 C (long term) Cells counted with standard hemocytometerchamber with Neubauer ruling. The cells in 9 large squares counted & thenmultiplied by 0.6 to get number of cells per cumm of CSF.
Normal counts Cattle, sheep and pig : 0 - 15 cells/ cu mm Dog : upto 25 cells/cu mm Horse : upto 23 cells/cu mm Increased number of white blood cells (pleocytosis) occurs ininflammatory lesions or irritation of brain and spinal cord.WBC > 200 cells/mlRBC > 400 cells/ml
• Marked increase – 100-500/μl– Acute pyogenic meningitis– Brain or spinal abscess• Moderate increase– Encephalitis• Mild increase– Neoplasia– Viral infections – 10-100(rabies – upto 500)Hemacytometer grid. The large cells withslightly irregular cell margins are WBCs. RBCsare smaller, light tan in color and round
Differential count Indicated when total count is elevated Various methods :Centrifugation – If the total cell count is less than 500 cells/μl. Membrane filtration – Even small Number of cells can beexamined. Sedimentation technique. For staining – Romanowsky stains Wrights Wright-Geimsa Leishman’s stain Also rapid staining methods : Diff-quik
• Neutrophils- Not seen in CSFoBacterial encephalitis/meningitisoAbscessoHemorrhage• LymphocytesoViral infectionsoAbscessoFungal infections – Cryptococcus neoformansoPost vaccinal inflammationoChronic conditions• Neoplastic cellsoLarge cells arranged in clusters
Myelin fragment in CSF from a horse withnecrotizing encephalomyelitis. Large sphericalstructure near macrophage contains a longspiral fragment of myelin. (Wright’s stain)
Lymphoma in the CSFMedium to large lymphocytes with immaturechromatin, prominent nucleoli and basophilic,vacuolated cytoplasm.(Wright- Giemsa)
Bacteriological examinationIt is carried out when theCSF cell count and proteincontents are high.The organisms are isolatedin CSF and identified bycultural methods.Organisms detected aretoxoplasmas,trypanosomes,bacteriaGram stained CSF showing gram positiveAnthrax bacilli