Inhibition Of Pathogenic Antibodies In Myasthenia Gravis To The Binding Of Acetylcholine Receptor By A Single Chain Antibody Fragment Fused With Human Serum Albumin

Objective: To investigate in vitro the therapeutic effects of single chain variable fragment of antibody fused human serum albumin 637 (HSA-ScFv637) on myasthenia gravis (MG). Methods: After resuscitating the Pichia pastoris GS115 strains which bear recombinant gene expression vector, the strong resistant G-418 of multiple copies of transformants were selected. A single colony was incubated in YPD, BMGY and BMMY culture media to induce expression in turn. Within 3 days of inducing expression, 100 % methanol was added to BMMY every 24 hours to make the final concentration of 1 % in total culture medium. After centrifuging the supernatant of expression medium at 14000 g at room temperature, the target protein in supernatant was detected primarily by dot blot. The molecular weight of expressed protein (HSA-ScFv637) was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. The inhibitive effects of HSA-ScFv637 on the binding of sera of MG patients were measured by competitive ELISA. Results: HSA-ScFv637 were able to inhibit the binding of the sera of MG patients to human AChR from 2.0% to 77.4%, and the inhibition ratio was decreased with decay of HSA-ScFv637. Conclusion: HSA-ScFv637 has in vitro a protective effect on human AChR from binding of the pathogenic anti-AChR antibodies to AChR.