Detection of Acetylcholine Receptor Blocking Antibodies by Flow Cytometry.

OBJECTIVES: Detection of acetylcholine receptor (AChR) blocking antibodies through the use of a radiolabel has become standard procedure in most laboratories. Known drawbacks associated with radioassay, including cost of radioisotopes, hazards to laboratory professionals, and manufacture and disposal of radioactive materials, have prompted investigation into replacement assays. We describe here a high-throughput immunofluorescent flow cytometric assay designed for the detection of AChR blocking antibodies. METHODS: In total, 323 serum samples were tested on both the AChR blocking radioassay and the new immunofluorescent flow cytometric assay. RESULTS: Analysis of the results revealed a 96.9% concordance between the two assay methods. CONCLUSIONS: Our results indicate that a new immunofluorescent flow cytometric AChR blocking antibody assay is not only feasible but clinically comparable in both sensitivity (91%) and specificity (99%) compared with radioassay.

APhL antibody ELISA as an alternative to anticardiolipin test for the diagnosis of antiphospholipid syndrome.

BACKGROUND: Persistent levels of antiphospholipid (aPL) antibodies [lupus anticoagulant (LA), anticardiolipin (aCL), anti-beta 2 glycoprotein I (aß(2)GPI) IgG and/or IgM] in association with clinical features of thrombosis and/or pregnancy associated morbidity are indicative of antiphospholipid syndrome (APS). Of the aPL antibodies, aCL is the most sensitive for APS, however, their lack of specificity constitute a laboratory and clinical challenge. IgG/IgM antibodies directed against APhL (a mixture of phospholipids) has been reported to predict APS more reliably than aCL tests. The main objective of this study was to evaluate the performance characteristics of the APhL IgG/IgM ELISA, relative to the aCL and aß(2)GPI tests. METHODS: Sixteen (16) clinically confirmed APS and 85 previously tested serum (PTS) samples for aCL and aß(2)GPI IgG/IgM antibodies were evaluated with the APhL IgG/IgM ELISA. Clinical specificity was determined in 100 serum samples (50 healthy and 50 infectious disease controls [parvo- and syphilis-IgG/IgM positive]). RESULTS: The IgG antibody prevalence for aCL and APhL in the APS and PST groups was comparable with marginal differences in clinical specificities. In contrast to the aCL IgM ELISA, the APhL test showed improved clinical specificities (72% aCL vs 94% APhL in the healthy controls; 38% aCL vs 78% APhL in the infectious disease controls) with implications for increased reliability in the diagnosis of APS. The overall agreement of the APhL with the aCL or aß(2)GPI for the IgG tests was 89% and 85% respectively, and that of the APhL IgM to the aCL or aß(2)GPI IgM tests was 72% and 86% respectively. CONCLUSION: Routine use of the APhL IgG/IgM ELISA may substantially reduce the high number of false positives associated with the aCL test without loss in sensitivity for APS.