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. Author manuscript; available in PMC: 2013 Jan 1.
Published in final edited form as: Ann Rheum Dis. 2011 Aug 21;71(1):92–98. doi: 10.1136/ard.2011.151712

Figure 3.

Figure 3

Beta and gamma actin are citrullinated autoantigens in RA. A. Beta- and gamma-actin are modified in ionomycin activated neutrophils and the novel species co-migrates with the 47kDa RA autoantigen. Samples from control (lanes 1, 3, and 5) and ionomycin activated neutrophils (lanes 2, 4 and 6) were analyzed by immunoblotting using monoclonal antibodies against human beta- and gamma-actin (Sigma) or RA serum. The filled arrow denotes the 47kDa species detected by anti-actin antibodies and RA sera. B, C. Beta- and gamma-actin are citrullinated and recognized by RA autoantibodies. Recombinant human beta-actin (lanes 1 and 2) or gamma-actin (lanes 3 and 4) was incubated in the absence (lanes 1 and 3) or presence (lanes 2 and 4) of rabbit PAD2. After 1 hr at 37°C, samples were divided in two and analyzed by electrophoresis on 8% SDS-polyacrylamide gels. After electrophoresis, proteins were visualized by Ponceau S staining (upper panels) prior to AMC immunoblotting (B, lower panel) or immunoblotting with RA sera (C, lower panel). Data from 1 representative serum containing anti-citrullinated actin antibodies is shown in C.