Comparison of monoclonal antibodies for flow cytometric analysis of HLA-B27 antigen

HLA-B27 is an antigen associated with the disease ankylosing spondylitis. Ninety percent of Caucasians with ankylosing spondylitis possess the HLA-B27 antigen. However, only 20% of Caucasians with the HLA-B27 antigen will develop the disease. Defining the presence or absence of the HLA-B27 antigen can be helpful in differentiating ankylosing spondylitis from juvenile rheumatoid arthritis. In this study, we evaluated the application of two monoclonal antibodies (MoAbs), using flow cytometric analysis for the detection of HLA-B27 antigen. After an initial comparison of HLA-B27 analysis by flow cytometry to the standard microlymphocytotoxicity assay, cutoffs were established to differentiate HLA-B27 positive from HLA-B27 negative. One MoAb showed very reliable results with > 99% accuracy in discriminating HLA-B27 positive from HLA-B27 negative samples. Various parameters were investigated to obtain the optimum results and showed that incubation time, reagent lot, and the flow cytometric instrument can affect the results. We concluded that a reliable MoAb and flow cytometry are valuable for the rapid and inexpensive determination of HLA-B27 typing in the clinical setting. However, testing conditions can affect the accuracy of results; therefore, adequate parallel testing in various conditions must be performed in order to establish the proper standards.