Development of a highly sensitive diagnostic assay for muscle-specific tyrosine kinase (MuSK) autoantibodies in myasthenia gravis.

Autoimmune myasthenia gravis is usually characterized by the presence of autoantibodies against the acetylcholine receptor (~80-90% of patients) or muscle-specific tyrosine kinase (MuSK) (~5% of patients). In the remaining patients, no such antibodies (Abs) are detectable, but this could be due either to the presence of auto-Abs to yet unidentified antigens or to concentrations of circulating Abs below the threshold of the available assays. The most popular and sensitive assay for anti-MuSK Abs is a radioimmunoprecipitation assay (RIPA), which uses (125)I-labeled MuSK as test antigen. A serious limiting factor of the sensitivity of such RIPAs is that small volumes of test serum are required (maximum 5-20 µl) in order to avoid excessive background values. We have overcome this obstacle by the development of a two-step RIPA. This involves non-stringent affinity purification of anti-MuSK Abs from a large volume of patient's serum, followed by a regular RIPA step, with the isolated Abs, to determine the antibody titer. This two-step assay was shown to be 10-50 times more sensitive than the regular RIPA. All tested sera previously found to be positive in the regular RIPA and normal sera were also positive or negative in the two-step RIPA, respectively. Importantly, of seven tested sera previously characterized by regular RIPA as negative with titers that were above zero, but statistically not significantly higher from the background, two were found to be positive, while the others were clearly shown to be negative. We conclude that our diagnostic test can detect very low concentrations of circulating anti-MuSK Abs. The general principle of this two-step RIPA approach may also be applied to the detection of currently undetectable concentrations of circulating auto-Abs involved in other diseases.

[Expert research on sera yielding false-positive results for HIV antibodies during screening]. / Ekspertnye issledovaniia syvorotok, daiushchikh lozhnopolozhitel'nye rezul'taty na antitela k VICh pri skrininge.

The significance of different serological methods and assay systems for the verification of false positive cases of HIV infection has been analyzed on the basis of materials obtained in arbitration studies. As demonstrated by this analysis, the use of such highly specific and sensitive systems as Huma-Lab, Enzygnost, Serodia and Erythrorecombinant has made it possible to obtain a reliable result as early as at the first stage of expert diagnosis in the enzyme immunoassay and the agglutination test. The methods of radioimmunoprecipitation and indirect immunofluorescence have permitted a more precise differentiation of doubtful results than that achieved by immune blotting.