Cerebrospinal fluid (CSF) samples (n=50) from patients with neurological disease (bacterial infection, viral infection, neuroborreliosis and multiple sclerosis) were analysed to characterize cell populations by fluorescent immunocytometry with the CD-Sapphire haematology analyser. Reagent combinations applied to all CSF samples comprised CD3/CD19/HLA-DR and CD4/CD8, with some being further analysed using CD3/CD4, CD3/CD16 and CD3/CD25 protocols. Of the 50 samples, 11 were excluded because of high proportions of nonviable cells (n=2) or insufficient cell numbers (n=9). Apart from bacterial infection with granulocytosis, all diagnostic groups showed high proportions (51.4-77.0%) of CD3+ T cells. There was a modest association between T-cell and B-cell counts, but absolute B-cell numbers exceeded 5 cells/microl in only 7/39 cases (neuroborreliosis, n=6; bacterial meningitis, n=1). CD3/Ia antigen (activation) co-expression was low and only exceeded 5% in 7/39 samples with no diagnostic correlation. Primary CD4+ and CD8+ T-cell subsets showed similar quantitative trends and CD4/CD8 co-analysis revealed the presence in all diagnostic groups (neuroborreliosis and multiple sclerosis in particular) of a CD4+CD8int fraction that was predominantly CD3+ and CD16- and had a morphological profile consistent with small lymphoid cells. Supplementary CD-Sapphire cellular immunological analysis of most CSF samples is feasible using the procedure detailed in this communication.