Cell analysis in cerebrospinal fluid (CSF) using Sysmex® hematology analyzers XT‐4000i and XE‐5000: Evaluation with CSF controls of the Joint German Society for Clinical Chemistry and Laboratory Medicine (DGKL)

Abstract

In cerebrospinal fluid (CSF) analysis, hematology analyzers (HAs) Sysmex® XT-4000i and XE-5000, equipped with flow cytometry (FCM), were used to count cells and differentiate leukocytes into mononuclear and polymorphonuclear cells (MNCs, PMCs) applying body fluid mode. FCM was evaluated with 20 DGKL CSF controls containing viable human leukocytes and erythrocytes. HA values were compared with reference values by Passing/Bablok regression analysis to reveal conformity. Conformity of white blood cells (WBCs) was obtained with native leukocytes, counted in calibrated Fuchs-Rosenthal chamber as reference; red blood cell counts proved inaccurate. CV <40% with WBC counts <20 per μL impairs accuracy. Reference WBC differentiation was assayed using FACS Canto II™ and FC-500 SN with anti-CD45, anti-CD14, anti-CD16, anti-CD16/56 [Becton Dickinson (BD); Beckman Coulter (BC)]. BD FACS lysing solution®-no-wash-procedure was applied. BC pretreatment with Versalyse lysing solution was not recommended. MNCs (lymphocytes + monocytes) were significantly lower (∼14%) on both HAs; PMCs (granulocytes or sum of neutrophils + eosinophils + basophils: range 1-86 M/L) were significantly higher (∼2.2-fold). WBC HA differentiation is not reliable because MNC/PMC differentiation yielded lower and higher values than FACS-FCM references, respectively. This is attributed to incorrect discrimination of leukocytes with rounded/nonrounded nuclei; adding leukocytes with nonrounded nuclei to too low HA MNCs (about 40% not-activated) yielded P/B conformity; subtraction of leukocytes with nonrounded nuclei from elevated HA PMCs showed conformity (about 85% activated). Nucleus/activation state of leukocytes was assessed using microhistology. Sysmex XT-4000i and XE-5000 HAs systems are inappropriate for complete CSF cell analysis.