Denielle

 Genesis  B.  Camato  

   IX.  CSF  ANALYSIS  
ANALYSIS  O F  URINALYSIS  AND  BODY  FLUIDS  |  REVIEWER  

FORMATION & PHYSIOLOGY TRAUMATIC COLLECTION

1. UNEVEN DISTRIBUTION OF BLOOD
† First recognized by COTUGNO, 1974 ê blood from intracranial haemorrhage will be
† Third major body fluid of the body evenly distributed throughout the three CSF
† Provides nutrients to the nervous tissue to remove specimen tubes whereas the traumatic tap
metabolic wastes will have the heaviest concentration ¢ tube 1,
† Produce mechanical barrier to cushion brain and gradually diminishing amounts in tube 2, and
spinal cord against trauma tube 3; traumatic procedure
MENINGES¢those that lined the brain and spinal cord
THREE LAYERS: 2. CLOT FORMATION
† Dura mater ê traumatic tap may form clots owing to the
† Arachnoid mater introduction of plasma fibrinogen into
† Pia mater specimen; intracranial haemorrhage will not
SUBARACHNOID SPACE- where CSF flows; located between contain fibrinogen to clot
arachnoid & pia mater
† Approximately 20mL /hr in choroid plexuses & TUBERCULAR MENINGITIS- classic web-like pellicle is seen
reabsorbed by the arachnoid villi to maintain total after overnight refrigeration of the fluid
volume of 140-170 mL in adults & 10-60mL in
neonates 3. XANTHOCHROMIC SUPERNATANT
BLOOD-BRAIN BARRIER- used to represent the control ê results of blood that has been present longer
and filtration of blood components to the CSF and then to than that introduced by the traumatic tap;
the brain care should be taken because a very recent
haemorrhage would produce a clear
SPECIMEN COLLECTION & HANDLING supernatant & introduction of serum protein
† CSF routinely collected by LUMBAR PUNCTURE from traumatic tap could also cause the fluid
between third, fourth, fifth lumbar vertebrae to appear xanthochromic.
† Specimen is usually collected in three sterile tubes
labelled 1, 2, 3 in the order to which they are Additional tests for differentiation: microscopic examination &
withdrawn D-dimer test
TUBE 1- CHEMICAL & SEROLOGICAL TESTS m Microscopic: macrophages ingesting RBCS
TUBE 2– MICROBIOLOGY (erythrophagocytosis or hemosiderin granules)=
TUBE 3- CELL COUNT- least likely to contain cells intracranial hemorrhage
introduced by spinal tap procedure (other methods:
Systernal and Ventricular puncture) m D-Dimer: detection of fibrin degradation product by
If specimen cannot be performed on STAT basis; specimens Dimer, latex aggln immunoassay indicates formation
should be maintained in the ffg manner: of fibrin at a haemorrhage site
† Hematology tubes are refrigerated
† Microbiology tubes remain at room temperature
† Chemistry and Serology tubes are frozen CELL COUNT

APPEARANCE † Routinely performed on CSF specimens is the WBC

† Normally crystal clear CSF count
† Crystal Clear, Cloudy/ Turbid, Milky, Xanthochromic, † RBC counts are usually determined only when a
hemolyzed/ Bloody traumatic tap has occurred and a correction for
† XANTHOCHROMIA WBC count is needed
ê term used to described CSF supernatant that † Cell count should be performed immediately because
is pink, orange or yellow- pink (very slight WBCs (particularly the granulocytes) and RBCs will
amount of oxyhemoglobin) to orange (heavy begin to lyse within 1 hour with 40% of leukocytes
hemolysis) to yellow (conversion of disintegrating after 2 hours
oxyhemoglobin to unconjugated bilirubin
† Other causes of xanthochromia: elevated serum
bilirubin, presence of pigment carotene, increase
protein concentrations and melanoma pigment
† Xanthochromia is due to immature liver function
seen commonly in infants particularly those
premature

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 Denielle  Genesis  B.  Camato  
   IX.  CSF  ANALYSIS  
ANALYSIS  O F  URINALYSIS  AND  BODY  FLUIDS  |  REVIEWER  

METHODOLOGY CORRECTIONS for CONTAMINATIONS

† Normal adult CSF contains 0-5 WBC/uL; higher in WBC (added)= WBC (blood) X RBC (CSF)
children (as many as 30 mononuclear cells/uL) RBC (blood)
consider as normal in newborns
† Improved Neubauer counting chamber is † When peripheral blood RBC and WBC counts are in the
traditionally used normal range, many laboratories choose to simply
† Calculation formula: subtract 1 WBC for every 700 RBCs present in the
Number of cells counted x dilution =cells.uL CSF
Number of squares x volume of 1 square
† Differential count should be performed on a stained
† This formula can be used in both diluted and
smear and not from the cells in the counting
undiluted specimens
chamber
† When performing diff count, 100 cells should be
counted, classified, and reported in terms of
percentage
† If the cell count is low and finding 100 cells is not
possible, report only the numbers of the cell types
seen

CYTOCENTRIFUGATION
† As little as 0.1mL of CSF combined with one drop of
30% albumin produces an adequate cell yield when
processed with the cytocentrifuged.
† Addition of albumin increases the cell yield and
TOTAL CELL COUNT decrease the cellular distortion frequently seen on
cytocentrifuged specimens
Clear specimens may be counted undiluted, provided
†
no overlapping of cells is seen during the microscopic
CSF CELLULAR CONSTIUTENTS
examination
† Adults have usually predominance of lymphocytes to
† When dilutions are required, calibrated automatic
monocytes (70:30) ratio whereas monocytes are
pipettes are used
more prevalent in children.
† Dilutions for total cell counts are made with normal
† Pleocytosis- increased numbers of these normal
saline mixed by inversion and loaded in the
cells is considered abnormal, as the finding of
hemocytometer by a Pasteur pipette
immature leukocytes, eosinophils, plasma cells and
† Cells are counted in the four corner square on both
macrophages, increased tissue cells & malignant cells
sides of the haemocytometer
† High WBC count of which the majority of the cells
Clarity Dilution Amount of Amount of
are neutrophils is indicative of bacterial meningitis
Sample Diluent
† High percentage of lymphocytes & monocytes
Slightly hazy 1 : 10 30μL 270 μL suggests: viral, tubercular, fungal, or parasitic origin
† Cell forms differing those found in blood include
Hazy 1 : 20 30μL 570 μL macrophages, choroid plexus, and ependymal cells,
Slightly 1 : 100 30μL 2970 μL spindle-shaped cells, and malignant cells
cloudy
Slightly 1 : 200 30μL 5970 μL
bloody
Cloudy 0.1 mL of a 9.9 mL
Bloody 1 : 10,000 1:100 dilution
Turbid
WHITE BLOOD CELL COUNT

† Specimens requiring dilution can be diluted,

substituting 3% acetic acid to lyse the RBCs
† Addition of methylene blue to the diluting fluid will
stain the WBCs providing better differentiation
† To prepare a clear specimen that does not require
dilution for counting, place four drops of mixed
specimen in a clean tube.
† Rinse a Pasteur pipette with glacial acetic acid,
draining thoroughly and draw the four drops of CSF
into the rinse pipette.
† Allow the pipette to sit for 1 minute, mix the solution
in the pipette, discard first drop and load the
hemocytometer

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 Denielle  Genesis  B.  Camato  
   IX.  CSF  ANALYSIS  
ANALYSIS  O F  URINALYSIS  AND  BODY  FLUIDS  |  REVIEWER  

CSF CELLULAR CONSTITUENTS † CSF – GLUCOSE

† Nucleated Red Blood Cells (NRBCs)- result of bone ê CSF glucose is 60-70% that of plasma glucose
marrow contamination during spinal tap ê Eg: Viral Meningitis ¢ slightly decrease glucose
† Eosinophils- Parasitic infection, fungal infection, ê Bacterial Meningitis ¢ decrease glucose
introduction of foreign material in CNS ê Blood glucose should be drawn in conjunction with
Eg: Rat Lung Worm (Angiostrongylus cantonesis) CSF values; blood glucose should be drawn 2
Entamoeba histolytica- causing amebic hours before the spinal tap
encephalitis
N. Fowleri, Acanthamoeba † CSF - LACTATE
† Macrophages- appear 2-4 hours after RBCs enter ê Levels greater than 35 mg/dL indicate bacterial
the CSF; indicative of previous haemorrhage meningitis thus increase in lactate
† Choroidal Cells- from epithelial lining of the choroid ê Viral Meningitis- produces lactate levels lower
plexuses than 25 mg/dL thus decrease in lactate
† Ependymal Cells- from linings of ventricles and neural ê Elevated in destruction of CNS tissue owing to
canal oxygen deprivation
† Spindle-shaped cells- from lining cells of arachnoid ê Falsely elevated results: bloody/hemolyzed/
† Malignant cells of hematologic origin (blasts from xanthochromic specimen
leukaemia, lymphoma cells)
† Malignant cells of non-hematologic origin (metastatic
carcinoma cells) eg: breast cancer † CSF - GLUTAMINE
ê produced from ammonia and alpha-
CSF ANALYSIS – CHEMICAL EXAMINATION ketoglutarate by the brain cells
ê serves to remove toxic waste product ammonia
† CSF - PROTEIN from the CNS
ê Most frequently performed chemical test on ê Normal levels of glutamine is between 8-18 mg/dL
CSF; Normally between 15-45 mg/dL ê Elevated levels are associated with liver
ê CSF protein is <1% compared to the total disorders/ hepatic coma & Reye’s Syndrome
serum protein
ê ALBUMIN is the major protein in the CSF CSF ANALYSIS - MICROBIOLOGIC EXAMINATION
ê PRE-ALBUMIN (Transthyretin) second ê Used to identify the cause of meningitis
most predominant protein in the CSF ê CSF CULTURE is confirmatory test
ê IgM, Fibrinogen, Beta-Lipoprotein are not ê Gram stain, acid fast stain, India ink and latex
found in normal CSF agglutination test serve for preliminary diagnosis
ê Tau transferrin is found only in CSF; not ê India Ink ¢ for Cryptococcus neoformans; look
found in blood for capsules (fungal meningitis)
ê Elevated total protein values are most ê Most frequent agents include: Streptococcus
frequently seen in pathologic conditions. pneumonia, Haemophilus influenza, Escherichia coli,
Abnormally low values are most frequently Strep agalactiae & Neisseria meningitis (all of
seen in pathologic conditions these are encapsulated organisms)
ê Methods of CSF protein measurement:
Turbidimetric Mtd (eg: SSA/TCA) Dye-binding CSF ANALYSIS - SEROLOGIC TESTS
Method (BCG/BCP) ê For assessment of neurosyphilis using VDRL or
ê CSF/Serum-Albumin index is used to assess FTS-Abs (Treponema pallidum)
the integrity of the blood brain barrier
ê IgG index used to assess if there is Formation and Composition of CSF
• Blood brain barrier maintains the relative homeostasis of CNS
intrathecal IgG production within the CSF
environment by tightly regulating the concentration of substances by
ê OLIGOCLONAL Bands- for assessment of
specific transport systems for H+, K+, Ca2+, Mg2+, HCO3-.
Multiple Sclerosis; these band are also present
• Glucose, urea and creatinine diffuse freely between blood and the CSF.
in disorders such as encephalitis, neurosyphilis,
• Proteins cross freely by passive diffusion along the concentration
Guillain-Barre syndrome and neoplastic gradient and is also influenced by molecular weight.
syndrome Composition of Normal CSF
ê Myelin-Basic Protein is indicative of recent Protein - 15 - 45 mg/dL
destruction of myelin sheath that protects Glucose - 50 - 80 mg/dL
the axon of neurons (demyelination) can be Urea - 6.0 - 16 mg/dL
used to monitor the course of multiple Uric acid - 0.5 - 3.0 mg/dL
sclerosis Creatinine - 0.6 - 1.2 mg/dL
Cholesterol - 0.2 - 0.6 mg/dL
Ammonia - 10 – 35 μg/dL
Sodium - 135 – 150 mEq/L
Potassium - 2.6 – 3.0 mEq/L
Chloride - 115 – 130 mEq/L
Magnesium - 2.4 – 3.0 mEq/L
Cells - 0 – 5 Lymph/μL
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