The document discusses biochemical analysis of cerebrospinal fluid (CSF) in medical laboratories. Routinely tested parameters in CSF include glucose, protein, electrolytes, lactate, and enzymes. Proper handling and centrifugation of CSF samples is important to avoid contamination. Abnormal levels of glucose, protein, lactate and enzymes can indicate conditions like meningitis or tumors. The Pandy's test and Biuret method are described for measuring CSF protein levels, along with normal ranges. Spectrophotometry is used to analyze results from colorimetric assays.

1. BIOCHEMICAL EXAMINATION
Hussein A. Abid
Laboratory Medicine Specialist
Iraqi Medical Laboratory Association
Scientific Affairs and Cultural Relations Section
Scientific Affairs and Training
Fourth lecture
26/09/2018 (Wednesday)
Dr. Maha Al-Saffar
Laboratory Medicine Specialist

2. CHEMICAL ANALYSIS
• Routinely performed biochemical tests in CSF
are:
1. Glucose
2. Protein (total and specific)
3. Urea and creatinine
4. Electrolytes (Na+, K+, Mg+2, Cl-)
5. Lactate and lactate dehydrogenase (LDH)
6. Glutamine and acid-base parameters.
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3. Remember !!
• Before analysis, the fluid should be centrifuged
to avoid contamination by cellular elements.
• CSF is the most precious biological material.
Often, only small volumes of CSF are available for
analysis due to difficulty in collection; hence
handle this with care.
• The specimen may contain virulent organisms,
so strict safety precautions should be followed.
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4. CSF GLUCOSE
• CSF glucose is normally approximately two-
thirds (2/3) of the fasting plasma glucose.
• A glucose level below 40 mg/ dL is significant
and occurs in bacterial and fungal meningitis
and in malignancy.
• Glucose is consumed by leukocytes and tumor
cells.
• It can be elevated in diabetes mellitus (DM).
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5. CSF PROTEIN
• Total protein levels in CSF are normally very low, and
albumin makes up approximately two thirds (2/3) of the
total. CSF protein may rise to 500 mg/dl in bacterial
meningitis.
• A more moderate increase (150-200 mg/ dL) occurs
in inflammatory diseases of meninges (meningitis,
encephalitis), intracranial tumors, subarachnoid
hemorrhage, and cerebral infarction.
• A more severe increase occurs in the Guillain-Barré
syndrome and acoustic and spinal schwannoma.
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6. CSF PROTEIN
• Multiple sclerosis (MS): CSF protein is normal or mildly
increased.
• Increased IgG in CSF, but not in serum (IgG/ albumin index
normally 10:1). About 90% of MS patients have oligoclonal IgG
bands in the CSF. Oligoclonal bands occur in the CSF only not in
the serum.
• The CSF in MS often contains myelin fragments and myelin basic
protein (MBP).
• MBP can be detected by radioimmunoassay. MBP is not specific
for MS. It can appear in any condition causing brain necrosis,
including infarcts.
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7. LACTATE & LDH
• Lactate: the CSF lactate is used mainly to help
differentiate bacterial and fungal meningitis,
which cause increased lactate, from viral
meningitis, which does not.
• Lactate dehydrogenase (LDH): this enzyme
is elevated in bacterial and fungal meningitis,
malignancy, and subarachnoid hemorrhage.
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8. PROTEIN TEST
Cerebrospinal Fluid (CSF) Processing In Medical Laboratory

9. PANDY’S TEST
Principle:
• Proteins are precipitated by a saturated solution of phenol in water.
• The reagent used is phenol (carbolic acid crystals dissolved in water) or,
pyrogallic acid or, cresol, usually termed as Pandy's reagent or Pandy's
solution.
Procedure
• One drop of CSF sample (collected from the patient by lumbar puncture
technique), is added to about 1ml of Pandy's solution. The turbid
appearance signifies the presence of elevated levels of globulin protein in
the CSF and is regarded as positive Pandy's reaction. The CSF from a
normal adult shows no turbidity or precipitates and this is a negative
Pandy's reaction.
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10. PANDY’S TEST
Reactions and results
• Proteins in the cerebrospinal fluid, normally albumin and globulin
are present in the ratio of 8 to 1. Increase in protein levels are of
diagnostic value in neurological diseases.
• The normal CSF is clear and transparent fluid. The Pandy's
reaction makes it translucent or opaque.
Positive test
• A bluish white streak of precipitated proteins is seen. The degree
of turbidity depends on the amount of protein in the CSF. It can
vary from faint turbidity (mild to moderate elevation in CSF
proteins) to dense milky precipitate (high protein content in CSF).
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11. CSF PROTEIN ASSAY
• Protein present in the CSF is detected by a commercial kits
based on Biuret method or any other method.
• Biuret reagent when interacts with the peptide bonds in the
protein give a blue colored product.
• The intensity of the colour is proportional the amount of protein
in CSF.
• Color intensity is determined by measuring the absorbance by
the colored solution at a wavelength of 546 nm.
• Absorbance is measured by an instrument, spectro-
photometer.
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12. SPECTROPHOTOMETER
Most of visible spectrophotometers are composed
of:
• Light source which works with visible wavelengths
(400-700 nm).
• Monochromator filter for choosing desired wavelength.
• Sample holder (cuvette).
• Detector.
• Meter or recorder.
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13. Cerebrospinal Fluid (CSF) Processing In Medical Laboratory
Spectrophotometer

14. PROCEDURE
• Mix and incubate for 15 minutes at room temperature. Measure
absorbance at 546 nm.
• Note: other kits based on another methods also could be used.
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15. CALCULATIONS
Protein (g/ L) =
Abs. of sample
Abs. of Std.
× Std. concentration
Since;-
• Abs. = absorbance
• Std. = standard
• Std. concentration depending on this kit is 60 (g/ L).
• Normal range: 15 – 45 mg/ dL = 0.15 – 0.45 g/ L
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16. OTHER PARAMETERS
• Other biochemical tests for CSF, will be as
protein test, are also depend on the
commercial kits.
• The biochemical testing results will help in
differentiating the types of meningitis.
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