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Cerebrospinal Fluid Cytology: a Highly Diagnostic Method for the Detection of Diseases of the Central Nervous System

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Cerebrospinal Fluid Cytology: a Highly Diagnostic Method for the Detection of Diseases of the Central Nervous System J Lab Med 2016; aop Mini Review Michael Torzewski* and Karl J. Lackner Cerebrospinal fluid cytology: a highly diagnostic method for the detection of diseases of the central nervous system DOI 10.1515/labmed-2016-0044 Received January 28, 2016; accepted March 10, 2016 Introduction Abstract: Cytologic examination of cerebrospinal fluid The cerebrospinal fluid (CSF) is a clear and colorless (CSF) is a technically simple, yet productive diagnostic extracellular aqueous liquid in the cavities of the central procedure. The cytocentrifuge technique is the most com- nervous system (CNS) (brain ventricles, central canal monly utilized method to concentrate the generally scant of the spinal cord) and the subarachnoid space in ver- cellular components of CSF. There are several preanalyti- tebrates. The subarachnoid space is a fluid-filled cavity cal and analytical pitfalls causing artefacts and making that covers the brain and spinal cord and communicates proper assessment of the CSF cell preparation more dif- with the ventricular system of the brain. It is delimited to ficult or even impossible. The common cell types of CSF the outside by the arachnoid membrane against the dura are lymphocytes and monocytes including their activated mater, and inwardly by the pia mater against the surface forms. Cytologic examination of inflammatory conditions of the brain. It is permeated with arachnoid trabeculae, puts emphasis on the cellular composition of CSF caused which contain loose connective tissue stroma. The arach- by bacterial infections compared to viral infections and noid membrane and pia mater form the leptomeninges. noninfectious inflammatory diseases of the brain. Con- The CSF protects the CNS against shock and pressure cerning non-neoplastic disorders, diagnosis of subarach- from the outside and also acts as a transport medium noidal hemorrhage is of special interest and a main field for a large number of soluble substances, such as elec- of application of CSF cytology. The cytology of neoplas- trolytes, glucose and immunoglobulins. It is formed in tic disorders encounters three typical constellations the the ventricles by specially differentiated epithelial cells investigator is usually confronted with: either a primary of the choroid plexus and the ependyma. As part of the malignancy is already known and dissemination to the CSF circulation, it drains from the ventricular system of meninges shall be evaluated or clinical and neuroradio- the base of the brain into the subarachnoid space. The logical findings are suggestive of neoplastic meningitis secretion rate is 500 mL/day. With a total volume of cer- though without sufficient evidence of the primary tumor. ebrospinal fluid of approximately 150–170 mL, this means And third, a spinal tap is performed for other reasons and that the entire CSF is renewed every 5–7 h [1]. CSF is gener- malignant cells are an incidental finding. ally obtained for diagnostic purposes by puncture of the spinal canal at LV 3/4 or 4/5. It may also be collected by Keywords: cerebrospinal fluid cytology; meningitis; neo- suboccipital puncture or ventricular drainage. plastic meningitis; subarachnoideal hemorrhage. Significance of practical *Correspondence: Prof. Dr. med. Michael Torzewski, MA, (conventional) CSF cytology in the Robert-Bosch-Krankenhaus GmbH, Abteilung für Labormedizin, overall spectrum of CSF diagnostics Auerbachstr. 110, 70376 Stuttgart, Germany, Tel.: 0711/8101-3501, Fax: 0711/8101-3618, E-Mail: [email protected] Karl J. Lackner: Universitätsmedizin Mainz, Institut für Klinische After the collection of 3–15 mL CSF by way of lumbar punc- Chemie und Laboratoriumsmedizin, Langenbeckstr. 1, Mainz, ture, the basic program of tests includes the analysis of Germany the CSF, as well as the determination of the cell count, 2 Torzewski and Lackner: Cerebrospinal fluid cytology glucose and lactate in the CSF. In addition, the albumin the fourth and fifth lumbar vertebrae. Where possible, and immunoglobulin quotients are analyzed and repre- sterile polypropylene tubes should be used as sample con- sented in what is known as a CSF/serum quotient diagram. tainers. Glass tubes cause cells to adhere to the container Without going into detail on these parameters, it should surface, thus falsifying the cell count [6]. EDTA or sodium be mentioned that they are mostly used in the diagnostics fluoride tubes are not suitable for a cytological diagnosis of inflammatory diseases affecting the CNS. either. A minimum of 2 mL CSF is required for cell counts In addition to determining the cell count of leukocytes and the production of cytological specimens. But as much and erythrocytes in the CSF, a differentiated cytological as 10 mL CSF may be recommended in connection with examination of at least one, or better yet, two specimens specific tests, such as for tumor cells. should be performed as a matter of principle. The wide- As CSF generally has only a low buffering capacity spread practice of creating cytology specimens without an as a result of its very small protein and cell contents, it explicit request on the part of the sender – if at all – only is crucial for the CSF to be transported to the laboratory above a certain number of cells can lead to fatal misin- and processed there immediately after being collected by terpretation with serious consequences for the patient. puncture. Otherwise, a cytological assessment will be ren- For example, tumor cells or parasites can be detected in dered difficult or, in most cases, impossible due to the pH specimens with normal or slightly elevated cell counts value that changes quickly from 7.32 to 7.36 (regular range (see below and [2, 3]). in CSF) to alkaline levels of 7.8 and higher, as well as due The cytological examination of cerebrospinal fluid is to the concomitant autolytic changes [7]. A duration of 2 h a technically simple procedure that is quick to implement. between the puncture and the production of the cytologi- It is cost-effective and, under diagnostic aspects, a very cal specimen is considered critical. If this cannot be done, productive method, but does require considerable expe- the CSF may be set with buffered formalin at a ratio of rience on the part of cytologists. Generally, CSF cytology 1:1. But this procedure should not be employed by way of plays a crucial role in the following [4, 5]: routine because of inevitable artifacts. – Detection and detailed mapping of pathogen-induced inflammatory diseases of the CNS – Detection of acute and past bleeding into the CSF space – Basic recognition and at least preliminary typing of Cell preparation (sedimentation) cells of malignant tumors with respect to their origin and staining of cell specimens CSF cytology is also often used to evaluate therapeutic The cell specimen for cytological diagnostics must be response, such as in the treatment of malignant tumors by produced within 2 h of the puncture. CSF cytology has means of chemotherapy. become a routine method at CSF laboratories thanks to As concerns the diagnostic significance of CSF cytol- the application of the Pappenheim panoptic staining ogy, the Reference Institute of Bioanalysis (RfB) conducted already commonly used for blood and bone-marrow cells a web-based interlaboratory test 2 years ago to enable the (combined from eosin-methylene blue staining according laboratory to perform regular checks on its own experi- to May-Grünwald and Azure-II-eosin staining according to ence in evaluating cytological CSF specimens and, thus, Giemsa; MGG staining) [4]. Special stains, such as Gram, to continuously improve the quality of CSF diagnostics. Prussian-blue and immunocytochemical staining, will be discussed in greater detail below. Pre-analysis in CSF cytology Cell populations of a normal and CSF punctures are difficult for patients, especially due activated CSF cell picture to the post-puncture low CSF headache syndrome (head- aches occurring within the first 5 days following a punc- A CSF sample obtained by lumbar puncture from a healthy ture), nor can they be repeated any number of times. patient generally contains two cell types, that is, lympho- Accordingly, particularly stringent requirements must cytes and monocytes. The ratio of lymphocytes to monocytes be in place for the careful processing of samples at the is around 70–30. As the puncture needle passes through pre-analysis stage. CSF is usually collected by means of various tissues on its way into the subarachnoid space lumbar puncture between the third and fourth or between (skin, fat and connective tissue, striated muscle), and also Torzewski and Lackner: Cerebrospinal fluid cytology 3 comes into contact with the cartilage and bones of the spine, Monocytes have a diameter of 15–20 µm, a lobed or cells of these tissues can be found in cytological specimens. indented nucleus and blue-gray cytoplasm. Occasionally, By the same token, a CSF sample taken from the ventricular smaller vacuoles are found there (Figure 1C). Many and/or system may contain choroid plexus epithelial cells, ependy- larger vacuoles are already an indication of an activated mal cells and fragments of cerebral parenchyma [5, 6]. state. In addition, activated monocytes, which are gen- Lymphocytes, at least in smaller quantities, are found erally also larger than non-activated forms, often have a in almost all CSF samples. These are small (diameter of 7–9 rounded nucleus (Figure 1D). µm), relatively isomorphic cells. The chromatin of the cell nucleus is usually dense and homogeneous. There is only a small cytoplasm edge that is stained blue, somewhat pale, Pathological CSF cell findings somewhat more intensively colored (Figure 1A, 1). Acti- vated lymphocytes are a form of differentiation between in connection with inflammatory normal lymphocytes and the plasma cell under inflam- diseases of the CNS matory conditions. Compared to non-activated lympho- cytes, they are larger (up to 25 µm in diameter) and have Inflammatory diseases of the CNS usually lead to sig- a wider cytoplasm edge as an expression of ribosomal nificant changes in the cellular composition of the CSF.
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