A human monoclonal autoantibody to a nucleolar structure

Abstract

SUMMARY Peripheral blood lymphocytes from a scleroderma patient (CDC) were isolated, transformed with Epstein-Barr virus and fused to the heteromyeloma SHM-D33. Supematants from cultures were screened for autoantibody production against nucleoprotamine by ELISA. Positive wells were cloned by limiting dilution. After cloning, supematants from two wells were positive for the nucleoprotamine assay. One named CDC-1 has been studied in our laboratory. CDC-1 recognized a nucleolar antigen by indirect immunofluorescence. By using an ELISA with purified recombinant antigens. CDC-1 reacted against Ro/SS-A, Ul (RNP) and Sm. By immunoblotting using a lysate of MOLT-4 cell line, CDC-1 was able to react against a structure of 60 k D. When the antigen recognized by CDC-1 was purified, SDS-PAGE under reducing conditions with purified antigen and subsequent silver staining of the gel allowed us to delect three bands at 60,55 and 39 kD, respectively. A screening by ELISA with previously characterized antisera against our purified antigen demonstrated reactivity of the CDC-1 antigen with those antisera able to recognize Ro/SS-A.