A standardized method of detecting antibodies to extractable nuclear antigens (RNP and Sm) by gel precipitation

Because of the importance of the detection of antibodies to RNP and Sm in the diagnosis of mixed connective tissue disease, systemic lupus erythematosus and certain forms of systemic sclerosis, the various factors which influence the sensitivity of the gel precipitation method for the detection of these antibodies were investigated. The agarose concentration, thickness, well sizes and distance between wells influence the sensitivity of the precipitin reactions. Suitable conditions for a sensitive and reproducible test system are specified. Because of variations in the antigenic composition of extractable nuclear antigen preparations made by different methods and from different tissue sources, a two-dimensional "chessboard titration" using serial twofold dilutions of a reference serum to test doubling dilutions of antigen preparations is recommended for standardizing different lots of antigen. Using the above-mentioned standardized agarose plates and appropriate reference serum with anti-Sm and/or anti-RNP antibodies in a chessboard titration, it is recommended that acceptable use dilution of antigen preparation be two doubling dilutions before reaching an endpoint. The reference sera selected should be compared to US reference standards supplied by the Center for Disease Control.