CSF

Formation

Mininges: 1. Dura mater 2. Arachnoid 3. Pia mater

Dura mater: outer layer

Arachnoid: filamentous (spider-like) inner membrane

Pia mater : lining surfaces of the brain and spinal cord

 Csf produced in choroid plexus: of 2 lumbar ventricles and 3 rd and 4th ventricles
 20ml per hour produced adults
 90 to 150 ml adults
 10 to 60ml neonates
 - arachnoid granulations / villae – reabsorbed – one way valve
 choriod plexuses are capillary networks that form the CSF from plasma by mechanisms of
selective filtration under hydrostatic pressure and active transport secretion
- tight junctures – prevents passage pf many molec – BBBd

Specimen handling and collection

 collected through lumbar puncture

 elevated pressure: req fluid withdrawn slowly
 spx collected 3 sterile tubes
- tube 1: chemical and serologic test- leasat affected by blood or bacteria
- tube 2: for microbio
- tube 3: cell count: least likely to contain cells by spinal tap

4th tube may be drawn : for better exclusion

EXCESS SHOULD BE FROZEN

Hematology tubes are refrigerated.

Microbiology tubes remain at room temperature.
Chemistry and serology tubes are frozen.

APPEARANCE
Crystal clear
- bedside
 Crystal clear, cloudy or turbid, milky, xanthochromic and hemolyzed/bloody

Cloudy, turbid, milky Increased protein or lipid conc, maybe

infection WBC
Xanthoxhromia - pink, orange or yellow
- presence pf rbc degradation products
- pink (oyhemoglobin)
- orange (heavy hemolysis
- yellow: unconjugated bilirubin
- others: carotene increased serum levels
, markedly increased protein conc,

Melanoma: meningeal melanosarcoma

-bilirubin due to immature liver fx in infants,

particularly in premature

Traumatic Collection (Tap

- due to intracranial hemorrhage, puncture of blood vessel

Uneven distribution of blood

 cerebral hemorrhage: evenly distributed in tubes
 traumatic tap: 1st tube heaviest conc, gradually decreasing

clot formation

- introduction of plasma fibrinogen into the spx

- Bloody CSF caused by intracranial hemorrhage does not contain enough fibrinogen to clot

Dse damage to bbb: increased filtration of protein and coagulatioan factors, no bloody fluid : meningitis,
froin syndrome and blockage of csf circulation

Classic weblike pellicle – tubercular meningitis, seen after overnight refrigeration of the fluid

Xanthochromic supernatant

- rbcs remain in csf 2hrs before noticeable hemolysis begins

- very recent hemorrhage would produce a clear supernatant, and introduction of serum protein from a
traumatic tap could also cause the fluid to appear xanthochromic

To examine a bloody fluid for the presence of xanthochromia, the fluid should be centrifuged in a
microhematocrit tube and the supernatant examined against a white background.

- differentiation includes microscopic examination and the D-dimer test

- macrophages containing ingested RBCs (erythrophagocytosis) or hemosiderin granules is indicative of

intracranial hemorrhage.

- Detection of the fibrin degradation product, D-dimer, by latex agglutination immunoassay indicates the
formation of fibrin at a hemorrha

CELL COUNT

 the leukocyte (WBC) count

 rbcs only when traumatic tap has occurred
 RBC COUNT : total cell count - wbc count
 - cell cpount should be performed immediately: wnc and rbc lyse within 1 hr, 40% leukocytes
disintegrating after 2hrs. apx not analyzed immediately refrigerated

METHODOLOGY

 Normal adult: 0 to 5 wbc

 Higher in children 30 mononuclear cells/ ul normal in newborn
 200wbc or 400 rbc/ul may be clear,necessary microscopic
 Improved Neubauer counting chamber for csf cell count

Calculation of csf count

# of cells counted x dilution = cells/uL

# of squares counted x volume of 1 sq

TOTAL CELL COUNT

- clear specimens may be counted undiluted, no overlapping pf cells is seen during the microscopic
examinarion

- dilution normal saline, inversion, loaded into the hemocytometer with Pasteur pipette

- counted in 4 corner sq and the center sq on both sides of hemocytometer

WBC COUNT

- lysis of rbc must be obtained prior to performing the wbc count on either diluted or undiluted spx

- substituting 3% glacial acetic acid to lyse the rbcs

Addition of methylene blue to the diluting fluid stains the wbcs, better differentiation bte neutrophils
and mononuclear cells

Clear spx: 4 drops of mixed spx in clean tube. Rinse a Pasteur pipette with 3% glacial acetic acid draining
thoroughly, draw 4 drops of csf into rised pipette.

- sit for 1min, mix, discard first drop, load to hemocytometer

If different # of sq. ic counted, std Neubauer formula should be used to obtain the # of cells per
microliter

Corrections for contamination

- to correct wbcs and protein artificially introduced to csf

Wbcs (added) = wbc (blood) x rbc (CSF)

Rbc (blood)

** subract 1 for every 700 rbcs present in csf

uality control of csf

Csf: normal- lymphocytes and mono

Neutrophil -bm, early v,f,t, p men

Pyknotic nuclei – degenerating cells, like ncrbs

*nrbcs- bone marrow from spinal tap

Lympo HIV and AIDS
Moderately elevated <50 wbcs ul - ms
Mono
Eosinophils - parasitic, fungal (coccidioides immitis)
Macrophage - previous hemorrhage
 Dark blue or black – hemosiderin
Yellow- hematoidin
nonpathological
* choroidal cells: EC of choroid p
*spindle-shaped cells – lining from arachnoid
Malignant cells of hematologic origin Lymphoblasts, myelo, mono
Lymphoma cells
Malignant cells nonhemato Metastatic carcinoma
Cns tumors, astrocytomas, retinoblastomas,
medulloblastomas

Protein found in CSF

found Not found

-major beta globulin: transferrin IgM
- tau Fibrinogen
- major gamma globulin: IgG, …IgA Beta lipoprotein