Cerebrospinal fluid (CSF) supplies nutrients to nervous tissue, removes metabolic waste, and produces a mechanical barrier to cushion the brain and spinal cord.
The fluid flows through the subarachnoid space located between the arachnoid and pia mater.
CSF is produced in the choroid plexuses of the two lumbar ventricles and the third and fourth venticles.
Appearance | Cause | Major Significance |
---|---|---|
crystal clear | normal | |
hazy, turbid, milky | microorganisms | meningitis |
protein | disorders of the blood-brain barrier | |
production of IgG within the CNS | ||
cloudy | WBCs | meningitis |
oily | radiographic contrast media | |
bloody | RBCs | hemorrhage |
traumatic tap | ||
xanthochromic | hemoglobin | old hemorrhage |
lysed cells from traumatic tap | ||
bilirubin | RBC degradation | |
increased serum bilirubin | ||
carotene | increased serum carotene | |
protein | disorders of the blood-brain barrier | |
melanin | meningeal melanosarcoma | |
clotted | protein | disorders of the blood-brain barrier |
clotting factors | introduced by traumatic tap | |
contains a pellicle | protein | disorders of the blood-brain barrier |
clotting factors | tubercular meningitis |
The cell count that is routinely performed on CSF specimens is the leukocyte (WBC) count.
Any cell count should be performed immediately, because WBCs (particularly granulocytes) and RBCs begin to lyse within 1 hour, with 40% of the leukocytes disintegrating after 2 hours.
A Neubauer counting chamber is routinely used for performing CSF cell counts.
They must be soaked in a bactericidal solution for at least 15 minutes and then thoroughly rinsed with water and cleaned with isopropyl alcohol.
Clear specimens may be counted undiluted, provided no overlapping of cells is seen during the microscopic examination.
Dilutions for total cell counts are made with normal saline mixed by inversion.
If only WBCs are needed, 3% glacial acetic acid is used for dilution to lyse the RBCs, along with methylene blue to stain and differentiate neutrophils and mononuclear cells.
Cells are counted in the four corner squares and the center square on both sides of the hemocytometer.
To correct for WBCs and protein artificially introduced into the CSF from a traumatic tap, the CSF RBC count and the blood RBC and WBC counts are needed.
The number of artificially added WBCs can be calculated using the following formula:
An approximate CSF WBC count can then be obtained by subtracting the “added” WBCs from the actual count.
The differential count should be performed on a stained smear and be different from the sample used in the counting chamber.
To ensure that the maximum number of cells are available for examination, the specimen should be concentrated prior to the preparation of the smear.
Type of Cell | Microscopic Findings | Major Clinical Significance |
---|---|---|
lymphocytes | all stages of development may be found | normal |
viral, tubercular and fungal meningitis | ||
multiple sclerosis | ||
neutrophils | disintegrate rapidly | bacterial meningitis |
early viral, tubercular and fungal meningitis | ||
cerebral hemorrhage | ||
monocytes | mixed with lymphocytes | normal |
viral, tubercular and fungal meningitis | ||
multiple sclerosis | ||
macrophages | may contain RBCs appearing as empty vacuoles or ghost cells, hemosiderin granules and hematoidin crystals | RBCs in spinal fluid |
radiographic contrast media | ||
blast forms | lymphoblasts, myeloblasts or monoblasts | acute leukemia |
lymphoma cells | resemble lymphocytes with cleft nuclei | disseminated lymphomas |
plasma cells | in traditional and classic forms | multiple sclerosis |
lymphocyte reactions | ||
ependymal, choroidal and spindle-shaped cells | in clusters with distinct nuclei and distinct cell walls | diagnostic procedures |
malignant cells | in clusters with fusing of cell borders and nuclei | metastatic carcinomas |
primary CNS carcinoma |
Adults usually have a predominance of lymphocytes to monocytes (70:30), whereas the ratio is reversed in children; occasionally, neutrophils are also present.
Following cytocentrifugation, neutrophils may also contain cytoplasmic vacuoles and have no granules.
Granules are also lost more rapidly in CSF.
Neutrophils with pyknotic or shrunken nucleii indicate degenerating cells and resemble nucleated red blood cells with multiple nucleii.
A high CSF WBC count with increased neutrophils is considered indicative of bacterial meningitis.
Increased neutrophils are also seen in the early stages (1 to 2 days) of viral, fungal, tubercular and parasitic meningitis.
Neutrophils associated with bacterial meningitis may contain phagocytized bacteria.
Neutrophils may also be increased following central nervous system (CNS) hemorrhage, repeated lumbar punctures, and injection of medications or radiographic dye; however, these are of little significance.
A moderately elevated CSF WBC count with a high percentage of lymphocytes and monocytes suggests meningitis of viral, tubercular, fungal, or parasitic origin.
Reactive lymphocytes containing increased dark blue cytoplasm and clumped chromatin are frequently present during viral infections.
Increased lymphocytes are seen in cases of both asymptomatic HIV infection and AIDS.
A moderately elevated WBC count (less than 50 WBCs/μL) with increased normal and reactive lymphocytes and plasma cells may be indicative of multiple sclerosis or other degenerative neurologic disorders.
Increased eosinophils are associated with parasitic infections, fungal infections (primarily Coccidioides immitis), and introduction of foreign material, including medications and shunts into the CNS.
Macrophages appear within 2 to 4 hours after RBCs enter the CSF and are frequently seen following repeated taps.
They tend to have more cytoplasm than monocytes in the peripheral blood.
The finding of increased macrophages is indicative of a previous hemorrhage.
Further degradation of the phagocytized RBCs results in the appearance of dark blue or black iron-containing hemosiderin granules.
Yellow hematoidin crystals represent further degeneration.
They are iron-free, consisting of hemoglobin and unconjugated bilirubin.
These cells are most frequently seen following diagnostic procedures such as pneumoencephalography and in fluid obtained from ventricular taps or during neurosurgery.
Choroidal cells are from the epithelial lining of the choroid plexus.
They are seen singularly and in clumps with distinct cell borders.
Nucleoli are usually absent and nuclei have a uniform appearance.
Ependymal cells are from the lining of the ventricles and neural canal.
They have less defined cell membranes and are frequently seen in clusters.
Nucleoli are often present.
Spindle-shaped cells represent lining cells from the arachnoid.
They are ususally seen in clusters and may be seen with systemic malignancies.
Lymphoblasts, myeloblasts and monoblasts in the CSF are frequently seen as a serious complication of acute leukemias.
Nucleoli are often more prominent than in blood smears.
Lymphoma cells are also seen in the CSF indicating dissemination from the lymphoid tissue.
They resemble large and small lymphocytes and usually appear in clusters of large, small, or mixed cells based on the classification of the lymphoma.
Nuclei may appear cleaved, and prominent nucleoli are present.
Metastatic carcinoma cells of nonhematologic origin are primarily from lung, breast, renal, and gastrointestinal malignancies.
Cells from primary CNS tumors include astrocytomas, retinoblastomas, and medulloblastomas.
They usually appear in clusters and must be distinguished from normal clusters of ependymal, choroid plexus, lymphoma, and leukemia cells.
Fusing of cell walls and nuclear irregularities and hyperchromatic nucleoli are seen in clusters of malignant cells.
The clinically important CSF chemicals are few, but under certain conditions, it may be necessary to measure a larger variety.
Normal values for total CSF protein are usually listed as 15 to 45 mg/dL, but are somewhat method dependent, and higher values are found in infants and older persons.
Elevated levels are caused by damage to the blood-brain barrier, production of immunoglobulins within the CNS, decreased clearance of normal protein from the fluid, degeneration of neural tissue, meningitis and hemorrhage conditions.
Decreased levels are caused by fluid leaking from the CNS.
Plasma protein can be artificially introduced into a specimen by a traumatic tap, so corrections to calculation must be made.
Both the cell count and the protein determination must be done on the same tube.
CSF proteins are measured using principles of turbidity production or dyebinding ability.
Albumin makes up the majority of CSF protein, while prealbumin is the second most prevalent fraction in CSF.
The major alpha globulins include haptoglobin and ceruloplasmin, the major beta globulin is transferrin; and the major gamma globulin is immunoglobulin G (IgG).
Only a small amount of IgA is present.
A separate carbohydratedeficient transferrin fraction, referred to as “tau,” is only present in CSF and used to confirm suspicious specimens collected.
Immunoglobulin M (IgM), fibrinogen, and beta lipoprotein are not present.
To accurately determine whether IgG is increased because it is being produced within the CNS or is elevated as the result of a defect in the blood-brain barrier, comparisons between serum and CSF levels of albumin and IgG must be made.
The CSF/serum albumin index evaluates the integrity of the blood-brain barrier.
An index value less than 9 represents an intact blood-brain barrier, and it increases relative to the amount of damage to the barrier
The CSF IgG index measures IgG synthesis within the CNS.
In general, values greater than 0.70 are indicative of IgG production within the CNS.
The CSF culture is actually a confirmatory rather than a diagnostic procedure; however, several methods are available to provide information for a preliminary diagnosis.
Specimens are concentrated by centrifugation at 1500 g for 5 minutes, because often only a few organisms are present at the onset of the disease.
However, at least a 10% chance exists that Gram stains and cultures will be negative; thus, blood cultures should be taken, because the causative organism is often present in both the CSF and the blood.
Bacteria commonly found such as Streptococcus pneumoniae, Haemophilus influenzae, Escherichia coli and Neisseria meningitidis can be detected using enzyme-linked immunosorbent assay (ELISA) or bacterial antigen test (BAT).
Streptococcus agalactiae and Listeria monocytogenes may also be encountered in newborns.
Acid-fast or fluorescent antibody stains are not routinely performed on specimens unless tubercular meningitis is suspected.
In cases of fungal meningitis, an India ink preparation is performed to detect the presence of thickly encapsulated Cryptococcus neoformans with a classic starburst pattern.
Latex agglutination tests may also be used but false-positive interferences may occur from interactions with rheumatoid factors.
Commercial kits that make use of incubation with dithiothreitol or pronase and boiling with ethylenediaminetetra-acetic acid can also be used with less false-positive interference.
bacterial | viral | tubercular | fungal |
---|---|---|---|
w/ neutrophils | w/ lymphocytes | w/ lymphocytes and monocytes | w/ lymphocytes and monocytes |
marked protein elevation | moderate protein elevation | moderate to marked protein elevation | moderate to marked protein elevation |
marked glucose decrease | normal glucose level | glucose decrease | normal to decreased glucose |
lactate >35 mg/dL | normal lactate level | lactate >25 mg/dL | lactate >35 mg/dL |
positive Gram stain and bacterial antigen tests | w/ pellicle | positive India ink and immunologic test for C. neoformans |