Objective To provide an alternative therapeutic modality for rheumatoid arthritis (RA), a novel bispecific antibody (BsAb) targeting human tumor necrosis factor alpha (TNF-alpha) and human complement component C5a was constructed. Results BsAb was expressed in Pichia pastoris and secreted into the culture medium as a functional protein. In vitro functional study demonstrated that BsAb could simultaneously bind to TNF-alpha and C5a and neutralize their biological actions. Furthermore, BsAb showed significant improvements in both the antigen-binding affinity and the neutralizing ability as compared to its original antibodies produced in E. coli. It was also found that TNF-alpha and C5a had an additive/synergistic effect on promoting the production of inflammatory cytokines and chemokines and C5a receptor (C5aR) expression in human macrophages. Compared to single inhibition of TNF-alpha or C5a with respective antibody, BsAb showed a superior efficacy in blocking inflammatory cytokines, chemokines, and C5aR response, as well as in lowering the C5a-mediated chemotaxis of macrophages via C5aR in vitro. Conclusions With improved production processing and the ability to simultaneously block TNF-alpha and C5a action, BsAb has a great potential to be developed into a therapeutic agent and may offer a better therapeutic index for RA.