Determination of anti-ADAMTS13 autoantibody titers in ELISA: Influence of ADAMTS13 presentation and autoantibody detection.

Background Immune-mediated thrombotic thrombocytopenic purpura (iTTP) is caused by inhibitory and/or clearing anti-ADAMTS13 autoantibodies. To determine the presence and total level of anti-ADAMTS13 autoantibodies, commercial and in-house developed enzyme-linked immunosorbent assays (ELISAs) are performed. However, different ELISA methods vary in relation to the presentation of recombinant (r)ADAMTS13 and the detection method of the anti-ADAMTS13 autoantibodies. Currently, the influence of those different approaches on anti-ADAMTS13 autoantibody titers is not known. Objectives To assess the influence of different ADAMTS13 presentation- and autoantibody detection methods on anti-ADAMTS13 autoantibody titers in ELISA. Materials/methods Anti-ADAMTS13 autoantibody titers from 18 iTTP patients were determined using four different set-ups of anti-ADAMTS13 autoantibody ELISAs. The ELISAs varied in the used presentation of rADAMTS13 (directly coated full-length rADAMTS13, directly coated rMDTCS and rT2C2 or antibody-captured full-length rADAMTS13) and the detection antibodies (polyclonal anti-human IgG or monoclonal anti-human IgG1-4 antibodies). Results Strong correlations between the different anti-ADAMTS13 autoantibody ELISA approaches were observed, when using polyclonal anti-human IgG detection antibodies recognizing all IgG subclasses similarly, independent of the method of rADAMTS13 presentation. Anti-ADAMTS13 autoantibody titers correlated less when using a mixture of monoclonal anti-human IgG1-4 , because not all IgG subclasses were recognized with similar affinities. Conclusion Anti-ADAMTS13 autoantibody levels using different methods of rADAMTS13 presentation strongly correlate. However, the levels of anti-ADAMTS13 autoantibodies are highly dependent on the used detection antibody, that should detect all IgG subclasses (IgG1-4) equally well.