Detection of autoantibodies in a quantitative immunoassay using recombinant ribonucleoprotein antigens

Abstract

A human cDNA expression library was screened with anti-ribonucleoprotein (RNP) antibodies from patients with connective tissue diseases. Three cDNA clones were isolated encoding 70 kD, A and B" ribonucleoprotein autoantigens which were expressed as beta-galactosidase fusion proteins. Antigens were purified and used to develop sensitive ELISAs suitable for the routine screening of large series of sera from patients with connective tissue diseases. More than 400 sera were tested both by ELISA and by immunoblotting. The ELISA was found to be at least as sensitive as immunoblotting and very specific. Anti-70 kD antibodies were found in 94% of patients with mixed connective tissue disease (MCTD), in 4% of patients with other connective tissue diseases but not in normal controls. Furthermore, the use of recombinant 70 kD antigen enabled us to discriminate between anti-70 kD antibodies present in anti-Sm and in anti-(U1) RNP sera. Recombinant A antigen contained at least two autoantibody-reactive sites; one unique for the A protein and another cross-reactive with anti-B" antibodies. Antibodies reactive with the unique site were found in 83% of MCTD patients, in 4% of patients with other connective tissue diseases and not in normal controls. Antibodies against the cross-reactive B" epitope present on A and B" recombinant antigens, were found in high titres in a small percentage of patients with systemic lupus erythematosus (SLE, 5%) and rheumatoid arthritis (RA, 2%).