EXAMINATION OF CSF

KAPIL GARG
BVSc & AH 2nd YEAR
B0044-2018
WHAT IS CSF ?

• Watery -  clear fluid, that is produced by central nervous system, the choroid
plexus inside the brain and circulates in brain and spinal cord.

• Found In subarachnoid space.

• This fluid is ultrafiltrate of plasma.

PROPERTIES  OF CSF

• Clear and colourless fluid.

• Contains glucose, electrolytes, amino acids, and less protein and few cells.

• It protects the CNS from injury, cushion it from the surrounding bone
structure.

• Removes waste products by returning them to the blood.

PURPOSE OF CSF ANALYSIS

• To diagnosis medical disorders that affect CNS - 

• Meningitis and encephalitis – viral, bacterial, fungal or parasitic

• Metastatic tumours and CNS tumours that shed cells into the CSF.
• Blood in the brain and spinal cord.
• Multiple sclerosis ( degenerative nerve disease )
METHODS OF COLLECTION OF CSF

• Can be collected from cisterna magna or subarachnoid space in the lumber

region.
 Cattle, sheep, goat - from suboccipital or lumber region
 Horse                            - from suboccipital puncture
Dog & cat                     - from suboccipital puncture
 Pig                                  - from only lumbar puncture

 5ml CSF  can be collected from large animal, 2-4 ml from dog and 10-30
drops from cat.
 Suboccipital puncture - 
• Site – anterior to the wings of atlas in the midline.
• Process - 
cast the animal on the right side and flex the head to left.
Shave, clean and sterile the area of puncture
Take a 3-4 inches long 16 gauge, sterile needle with a stylet and insert it at
the cervical midline at the level of cranial edge of the wings of atlas.
Direction of needle should be parallel to the long axis of the head.
When the needle enters the subarachnoid space you suddenly feel no
resistance.
Remove the stylet, CSF will flow out.
 Lumbar puncture - 
• Site – depression in the midline between the dorsal spinous processes of the
last lumbar vertebrae and the cranial edge of the median sacral crest.
• Process -
 Shave, clean and sterile the area.
Insert a 5 inches, 14 gauge needle with stylet in the midline.
On entering into the subarachnoid space CSF will flow out when the stylet is
withdrawn.
HOW TO PRESERVE THE CSF ?

• Cerebrospinal fluid should be examined soon after collection since

leukocytes degenerate rapidly.

• In case of delay, an anticoagulant should be added and it glucose is to be

estimated, fluoride is added.
SAMPLING

• Collected CSF sample is immediately divided into four tubes :-

Tube 1 : chemical investigation

Tube 2 : microbiology investigation
Tube 3 : microscopic examination
Tube 4 : cytology
EXAMINATION

• Examination of CSF is based on three types of examination :

 Physical examination

Chemical examination

Cytological examination
PHYSICAL EXAMINATION

 Colour – normally colourless like distilled water.

 If  it is dull red or brown in colour  then  it shows a haemorrhagic condition.
 If it is yellow then it shows icterus condition.
If it is green  then it shows suppurative conditions.

Turbidity – normally CSF is transparent.

 Note – turbidity does not become grossly evident until there are 500 or
more number of cells/ ul.
• Coagulation – normally CSF does not coagulate.
If fluid contains abnormal amounts of proteins, especially fibrinogen, it may
coagulate.
Internal haemorrhage or contamination during collection also cause its
coagulation.
Heavy coagulation in acute suppurative meningitis.

• Specific gravity – though its result are not considered very precisely useful in
the investigation of  neurological disease.
CHEMICAL EXAMINATION

• Protein – normal range is 12 – 40 mg/dl.

 protein concentration is increased in inflammatory and non-inflammatory
conditions.
 a crude test to find out the excess of protein is foam test.
 foam test – in this test, foams formed after shaking the CSF & will
disappear within a few minutes if it is normal.
Presence of foam even after 5 minutes indicates the presence of excess
proteins.
• Test for globulin – there are two test available :-
(1). Pandy’s test – Pandy's reagent(10 g of carbolic acid in 100 ml of distilled
water ) used I this test.
Procedure – take 1 ml of the saturated solution of Pandy's reagent and few
drops of CSF and shake the mixture.
- If turbidity develops then it means globulin is present.
- Result should be graded from + to +++ depending on turbidity.
(2). Nonne – apett ( Rass-jones) test – in this test take 1 ml of filtrated
saturated aqueous solution of ammonium sulphate. And carefully add 1 ml of
the CSF. Keep for few minutes.
- Interpretation – development of a white ring at the junction of two liquids
indicates the presence of globulin.

- Increased globulin in CSF indicates the presence of inflammation in CNS,

increase in the capillary permeability, hemorrhages or tissue destruction
occasionally resulting congestion, hemorrhage or edema.
• Glucose – concentration of glucose in CSF is approximately 60-70% of the
blood glucose level. In normal animals CSF glucose values range from 40-80
mg/dl.

Increased glucose levels (hyperglycorrhachia) occurs in hyperglycaemia,

encephalitis, compression of spinal cord, brain tumours and abscess of
brain, whereas decreased glucose level (hypoglycorrhachia) occurs in
hypoglycaemia and acute pyogenic infections.
CYTOLOGICAL EXAMINATION

• The total cell count in the CSF must be estimated within 20 minutes of
collection, since cells degenerate rapidly.
• Normal fluid contains small number of lymphocytes and rarely, histiocytes
and endothelial cells.
1. WBC count
2. Differential count
3. Bacteriological examination
(1). WBC Count –
Material – diluting fluid = crystal violet(0.1 ml) + glacial acetic acid(10 ml) +
distilled water ( 90 ml).
Procedure –
• Use white cell pipette of haemocytometer.
• Draw the diluting fluid up to 1 mark.
• Draw the CSF up to 11 mark.
• Mix thoroughly.
• Discard the first 2 or 3 drops.
• Charge the chambers on both sides of the standard haemocytometer.
• Wait for 2 minutes for the cells to settle.
• Count the cells on all squares on the 2 sides (that is on 18 sq. mm of the
area) Multiply the total number obtained by 0.6 to give the number of cells
in a µl of the CSF.
 normal count –
Cattle, sheep and Pig – 0 to 15 cells/µl
Dog – Up to 25 cells/µl
Horse – Up to 23 cells/µl
(2). Differential count - If the total count is less than 500 cells/µl, centrifuge
the fluid and make a smear of the sediment. Dry in air, stain with Leishman’s
stain and examine. Normally only lymphocytes or occasionally a few
mononuclear cells or histocytes may be found. Presence of neutrophils
indicates suppurative conditions.

(3). Bacteriological examination - Increase in lymphocytes than the normal

value indicates viral infection, fungal infection and toxic conditions. If the cell
count and the protein content are high, the aseptically collected CSF must be
subjected to various microbiological studies to ascertain the etiologic agent.
THANK YOU