Background: Enumeration of CD4+ and CD8+ T-cell subsets provides relevant information for diagnosis and monitoring of patients with cellular immunodeficiencies. As a result, an external quality assurance scheme was implemented in Belgium, The Netherlands, and Luxemburg in 1995. A workshop was held to train the participants in state-of-the art technology for assessment of absolute T-cell subset counts (i.e., a three or four-color, single-platform assay with lymphocyte gating based on CD45 and sideward light scatter) with the aim to achieve between-site coefficients of variation (CVs) <10% and within-site CVs <5% for ≥75% of the participants. Methods: Three send-outs of stabilized blood from a healthy donor were distributed to 55 laboratories, each with the request to perform the standard assay on three occasions. For comparison, each laboratory performed its local technique in parallel. Results: With the standard technique, between-site CVs of ∼8% (CD3+ T cells), ∼9% (CD4+ T cells), and ∼10% (CD8+ T cells) were achieved. Within-site CVs were <5% for 82% (CD3+ T cells) and ∼70% (CD4+ and CD8+ subsets) of the participants. Local techniques yielded between-site CVs of 13%-17% for CD3+, CD4+, and CD8+ T cells. Conclusions: The state-of-the-art technology for T-cell subset enumeration was implemented successfully among 55 Belgian-Dutch laboratories and resulted in significant reductions of between-site variation of absolute CD3+, CD4+, and CD8+ T-cell counts.

Flow cytometry, Lymphocyte immunophenotyping, Multicenter study, Standardization, Variation,
Department of Medical Oncology

Gratama, J.W, Kraan, J, Keeney, M, Granger, V, & Barnett, D. (2002). Reduction of variation in T-cell subset enumeration among 55 laboratories using single-platform, three or four-color flow cytometry based on CD45 and SSC-based gating of lymphocytes. Cytometry, 50(2), 92–101. doi:10.1002/cyto.10084