Diagnostic approach of phospholipid-dependent antibodies - State-of-the art lecture

Recent scientific developments enable to better understand the strong heter ogeneity of assay methods for phospholipid (PLP)-dependent antibodies, meas ured as lupus anticoagulant (LA) or as anti-PLP antibodies (APA). These lat ter are actually targeted at complexes of beta(2)-glycoprotein I (beta(2)GP I) and anionic PLP and also react with insolubilized beta 2GPI alone in som e patients. Although APA present little species specificity for beta 2GPI, some of them bind preferentially to complexes of human beta 2GPI and PLP. L As are diagnosed with screening assays (dPT, dRVVT, KCT, APTT); their sensi tivity is dependent on the concentration and type of PLPs used. They are ei ther beta 2GPI or prothrombin dependent. Based on the antibody-neutralizing capacity of PLP preparations (such as hexagonal phase phosphatidyletha-nol amine) or their bypassing activity, confirmatory assays enable to confirm L A. LAs and APAs bind to different epitopes on beta 2GPI or its complexes wi th PLP. Major characteristics of APA assays are: the capture antigen (PLPs, beta 2GPI source); its optimization and density on micro-ELISA plates; the efficacy of post-coating saturation; the animal serum used for saturation and diluent; the second antibody which must avoid nonspecific interactions; the calibrators proposed and their reference to international standards; t he cutoff between the normal and the pathological ranges; the assay sensiti vity and overall specificity for APAs. New optimized assays have been devel oped to meet these criteria. They are designed with PLP-coated plates, satu rated first with immunoglobulin-free human serum as a source of beta 2GPI, then with goat serum also used in diluent. The cutoff value is determined w ith at least 200 normal plasma samples (mean + 3 standard deviations or 99t h percentiles) and plasma samples from patients without APAs. This approach offers both optimized specificity and sensitivity for testing APAs, along with a good standardization and it helps to measure the true APAs associate d with pathology. Copyright (C) 1999 S. Karger AG, Basel.