External quality assessment of flow cytometric HLA-B27 typing

A biannual external quality assurance (EQA) scheme for flow cytometric typi ng of the HLA-B27 antigen is operational in The Netherlands and Belgium sin ce 1995. We report here on the results of the first seven send-outs to whic h 36 to 47 laboratories participated. With the send-out, four specimens fro m blood bank donors, who had been typed for HLA Class I antigens by complem ent-dependent cytotoxicity, were distributed. Subtyping of the HLA-B27 alle le was performed by PCR-SSP. Ten samples were HLA-B27(pos) (all HLA-B*2705) and 18 were HLA-B27(neg). For flow cytometry, the most widely monoclonal a ntibody (MoAb) used was FD705, followed by GS145.2 and ABC-m3. The majority of laboratories used more than 1 anti-HLA-B27 MoAb for typing. The HLA-B27 (pos) samples were correctly classified as positive by the large majority o f participants (median 95%; range 85% to 100% per send out); some participa nts considered further typing necessary and misclassification as negative w as only sporadically seen. The classification of HLA-B27(neg) samples as ne gative was less straightforward. Ten samples were correctly classified as s uch by 97% (82% to 100%) of the participants, whereas 64% (range 53% to 70% ) of the participants classified the remaining eight samples as HLA-B27(neg ). There was no significant prevalence of a particular HLA-B allele among t hese eight "poor concordancy" samples as compared to the ten "good concorda ncy" samples. Inspection of the reactivity patterns of the individual MoAb with HLA-B27(neg) samples revealed that ABC-m3 showed very little cross-rea ctivity apart from its well-known cross-reactivity with HLA-B7, whereas the cross-reactivity patterns of GS145.2 and FD705 were more extensive. The sm all sample size (n = 18) and the distribution of HLA-B alleles other than H LA-B27 did not allow assignment of specificities to these cross-reactions. Finally, we showed that standardized interpretation of the combined results of two anti-HLA-B27 MoAb reduced the frequency of false-positive conclusio ns on HLA-B27(neg) samples. In this series, the lowest frequency of false-p ositive assignments was observed with the combination of the FD705 and ABC- m3 MoAb. Cytometry (Comm. Clin. Cytometry) 42:95-105, 2000. (C) 2000 Wiley- Liss, Inc.