[Haematology] Cerebrospinal Fluid Analysis - Laboratory Examination

Cerebrospinal Fluid Analysis - Laboratory Examination, Cerebrospinal Fluid Analysis, Laboratory Examination, Graff's Textbook of Urinalysis and Body Fluids

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1. [Haematology] Cerebrospinal Fluid Analysis - Specimen Collection

Nearly every section of the laboratory can be involved in the evaluation of cerebrospinal ﬂuid. Laboratory tests that may be performed on CSF include, but are not limited to, macroscopic evaluation, microscopic evaluation of cell count and type, chemical analysis, microbiology cultures, and immunologic and molecular analyses. Some of these tests are beyond the scope of this text. However, the more common laboratory procedures are included.

PHYSICAL CHARACTERISTICS

Color and turbidity are noted. If a fourth tube is collected, or when the cell counts are completed, the tube may be refrigerated and observed for pellicle formation. Normal CSF is clear and colorless and demonstrates aviscosity similar to water. Abnormal turbidity is observed if blood cells, microorganisms, or flecks of protein are present. Varying degrees of CSF cloudiness due to the presence of cells is termed pleocytosis. An oily appearing CSF may contain radiographic contrast media.

Abnormal colors reflect the presence of various substances. Red blood cells can add a red, pink, or smoky color to CSF. If hemoglobin is present, the CSF can appear red or xanthochromic if the hemorrhage is older. Other substances that make CSF xanthochromic include bilirubin,

carotene, and melanin.

Because blood cells may be introduced into a CSF specimen at the time of lumbar puncture, careful differentiation between traumatic tap and hemorrhage must be made.

If there is a significant difference in the amount of blood present between the first and last tubes collected (later tubes gradually clearing), then the puncture was traumatic.

If all tubes collected show the same degree of blood, then a subarachnoid hemorrhage is most likely. Figure 9-5 demonstrates the difference in appearance of normal clear CSF, red CSF in hemorrhage, xanthochromic CSF from an old hemorrhage, and CSF from a traumatic tap.

If lumbar puncture is performed within the first 4 hours after a subarachnoid hemorrhage, the CSF will appear pale pink to red, depending on the degree of hemorrhage. Red blood cells lyse in CSF due to the low level of proteins and

Figure 9-5. Comparison of cerebral spinal ﬂuid appearance

between (A) normal CSF, (B) red CSF from fresh hemorrhage,

and (D) CSF from a traumatic tap

lipids as compared to plasma. After hemolysis, the CSF will change from a cloudy or hazy pink–red to a clear pink–red and then through various shades of light orange, yellow, and amber (xanthochromia), as oxyhemoglobin changes to methemoglobin, and then after about 12 hours bilirubin is formed. Gradual decrease in CSF color occurs over the ﬁrst 2 days, clearing in about 2–4 weeks.

Clotting of CSF is associated with a traumatic tap rather than hemorrhage, because CSF contains nearly no ﬁb- rinogen. Clotting of CSF may be present in cases of neu- rosyphilis and tubercular meningitis.

MICROSCOPIC EXAMINATION

Microscopic examination of CSF includes the counting of cells on a hemocytometer and differentiation of cell types on stained smears, as described in Chapter. As for all body ﬂuids, cell counts on CSF must be performed as soon as possible, because deterioration of cells in body ﬂuid speci- mens begins within 2 hours of collection. Cell counts are performed manually rather than using automation because of the low level of cells normally present.

CELL COUNTS

CSF hemocytometer cell counts are performed on wellmixed, undiluted specimens. However, if the CSF is grossly bloody, a dilution with saline may be necessary. Normally, no red blood cells are present in CSF. Red blood cells add little diagnostic value to CSF results but are often reported as they may help identify a traumatic tap.

Nucleated cells are counted as described in Chapter 7. A dilution with HCl eliminates erythrocytes and enhances nuclei. Some laboratories include the performance of a preliminary differential as part of their cell count. Nucleated cells may be classiﬁed as mononuclear or polymorphonu- clear. Although the nucleated cell count is reported as a leukocyte count, not all nucleated cells found in CSF are white blood cells (WBCs). Occasionally, ependymal cells or choroid plexus cells enter the CSF. Tumor cells may be pres- ent. Nucleated red blood cells may be present in specimens from traumatic taps during which a vertebral process was nicked.

Normal adult CSF can contain 0–5 WBCs per microliter. Children can exhibit higher CSF WBC counts; however nor- mal ranges are poorly documented.

DIFFERENTIAL COUNT

A CSF differential count is usually performed on cytocentrifuged preparations that have been stained with Wright stain. Of the few cells normally present in CSF, lymphocytes and monocytes are predominant. Neutrophils are not a common finding in CSF, and CNS lining cells are only rarely seen. Figure 9-6 pictures cells that can normally be found in CSF. In adults, normal proportions of cells in CSF usually range 28–96% lymphocytes, 16–56% monocytes, and 0–7% neutrophils. Eosinophils, ependymal cells, and histocytes are only rarely seen.

PLEOCYTOSIS

Pleocytosis is the term given to an increased amount of WBCs in a body fluid. CSF normally contains very few WBCs. The type of WBC present in CSF correlates with various forms of inflammation, infection, or malignant condition. WBCs that can be present in CSF include neutrophils, lymphocytes, plasma cells, eosinophils, monocytes, and macrophages. Other cells that may be present in CSF include CNS lining cells and malignant cells.

Figure 9-6. Cells that can normally be found in cerebral spinal ﬂuid.

(A) Neutrophil, (B) monocyte, and (C) lymphocytes.

Red blood cells are not normally present in CSF (Wright stain 1000x).

Figure 9-7. Neutrophilic pleocytosis in cerebral spinal ﬂuid (Wright stain 1000x).

Neutrophils

Neutrophilic pleocytosis is present in cases of bacterial meningitis and in the early stages of other forms of meningitis. Other causes for neutrophilic pleocytosis in CSF include cerebral abscess, subdural empyema, CNS hemorrhage, intrathecal treatments, and postseizure. Figure 9-7 illustrates neutrophilic pleocytosis. Causes for CSF neutrophilic pleocytosis are outlined in Table 9-1.

Lymphocytes

Lymphocytic pleocytosis predominates the later stages of meningitis that are viral, tubercular, fungal, or syphilitic in nature. Lymphocytes in CSF undergo the same morphologic changes as in peripheral blood, lending to the presence of various lymphocyte forms. Increased numbers of lymphocytes can also be seen in other inflammatory processes and degenerative disorders such as Guillian–Barré syndrome. Figure 9-8 illustrates lymphocytic pleocytosis. Causes for CSF lymphocytic pleocytosis are outlined in Table 9-1.

Figure 9-8. Lymphocytic pleocytosis in cerebral spinal ﬂuid (Wright stain 1000x).

Figure 9-9. Plasma cells in cerebral spinal ﬂuid (Wright stain 1000x).

Plasmacytes

Plasmacytes are not normally found in normal CSF. They can appear in the same disorders in which there is lympho- cytic pleocytosis. In addition, plasma cells can be seen in multiple sclerosis, where they may be the only abnormality. Figure 9-9 illustrates plasma cells in CSF. Causes for CSF plasmacytosis are outlined in Table 9-1.

Eosinophils

Eosinophils are a rare ﬁnding in normal CSF. If eosinophils comprise greater than 10% of cells in CSF, an eosinophilic pleocytosis is present. Eosinophils can be increased in para- sitic and fungal infections of the CSF or allergic reactions to malfunctioning intracranial shunts, radiographic contrast media, and drugs. Figure 9-10 illustrates eosinophils cells in CSF. Causes for CSF eosinophils are outlined in Table 9-1.

Monocytes and Macrophages

Monocytic pleocytosis is a rare ﬁnding. Although mono- cytes may be increased in CSF, they usually do not pre- dominate. Increased numbers of monocytes in CSF occur with increased numbers of other cells, mixed pleocytosis. Mixed pleocytosis can be present in chronic bacterial

Figure 9-10. Eosinophils in cerebral spinal ﬂuid (Wright stain 1000x)

Figure 9-11. Cerebral spinal ﬂuid with mixed pleocytosis (Wright stain 1000x).

meningitis, meningitis of tubercular or fungal origin, or rupture of a cerebral abscess. Figure 9-11 illustrates a mixed cell pleocytosis in CSF.

Macrophages originate from monocytes and are not a normal ﬁnding in CSF. Macrophages are a common ﬁnding after CNS hemorrhage and may be seen with phagocytized erythrocytes, digested erythrocytes, and hemosiderin (siderophages) or hematin crystals following decomposition of large amounts of hemoglobin. Macrophages that are present after CNS hemorrhage can help roughly identify the time at which hemorrhage occurred. Table 9-2 displays the changes that occur in the types of cells present after hemorrhage. A lipophage, macrophage with ingested fat, may be seen in CSF following brain infarct. Figures 9-12–9-16 illustrate macrophages and various inclusions.

Figure 9-12. Macrophage in cerebral spinal ﬂuid (Wright stain 1000x)

Other Cells

Other cells that may occasionally be present in normal CSF include ependymal cells, choroidal cells, and PAM cells. Ependymal cells can be seen in Figure 9-17. The morphology of these cells was discussed earlier in this chapter. Neonates normally can have increased numbers of these cells. Children with hydrocephalus will also have increased numbers of ependymal cells in their CSF. Ependymal cells and choroidal cells may be present in CSF in high numbers after traumatic brain injury, pneumoencephalography, surgery, myelography, ischemic infarction of the brain, ventricular shunts, and intrathecal injections. Nucleated red blood cells may also be present in specimens from a traumatic tap in which the vertebrae was nicked. Figure 9-18 shows the appearance of nucleated red blood cells in CSF. Ependymal cells, choroidal cells, and PAM cells can occur in clusters, making them difﬁcult to differentiate from clustered malignant cells. Malignant cells arise from various tumors, either primary CNS tumors or one that has metastasized to the CNS. Tumors that commonly metasta- size the CNS include carcinomas of the breast, gastrointestinal tract, lung, and leukemia and melanoma. Clumped CSF choroid plexus cells are seen in Figure 9-19.