Heterogeneous Nuclear Ribonucleoprotein A2 (of Hnrnp A2) And Anti-of Hnrnp A2/ra33 Antibodies In The Pathogenesis And Clinical Diagnosis Of Rheumatoid Arthritis Research

Heterogeneous nuclear ribonucleoprotein is the important nucleic acid-binding protein in RNA processing. In addition, a growing body of evidence points to hnRNP as an important target of the autoimmune response in rheumatic diseases and disease specificity in anti-hnRNP autoantibodies does exist. In fact, anti-hnRNP autoantibody in RA mainly directs to the hnRNP A2 protein, which is also termed anti-RA33 antibody. Anti-RA33/hnRNP A2 antibody is not only valuable diagnostic marker but may also allow additional insights into the pathogenesis of rheumatic diseases.In the study, the RA33 antigen were extracted and partial-purified from the nuclear of Ehrlich cells by affinity chromatography on heparin-sepharose CL-6B. SDS-PAGE showed: The bands of partial-purified antigen were obviously less than those of crude antigen, and main band existed in 33kD; The partial-purified antigen specifically reacted with the positive serum of anti-RA33 antibody by Western blotting. In order to further studies, monoclonal antibodies were produced from Balb/c mice immunized with the partial-purified antigen by applying hybridoma techniques. ELISA and Western blotting were applied to identify the positive clones and three hybridoma cell lines which secreted anti-RA33 McAb of IgG1 type were got. The McAb's characters were further analysed.Since anti-RA33 antibody mainly targets hnRNP A2, so we cloned hnRNP A2 gene, expressed and purified hnRNP A2 protein by genetic engineering techniques. First, total RNA was isolated from peripheral blood mononuclear cells and the RT-PCR was performed for acquiring hnRNP A2 cDNA. The fragment was cloned into PUC-T1 plasmids and identified by DNA sequencing. Second, the expression vector pET-28a-hnRNP A2 was constructed and transformed into E.coli BL21(DE3)plys S which...