Part Of CDNA Cloning, Expression And Polyclonal Antibody Of Complement C3from Lampetra Japonica

Complement C3is a member of immunology molecules, which is very important. In immune system, there are three complement activation pathways in mammals, alternative, lectin and classical pathways. Complement C3is works as the vital intersection molecule in these three pathways. With stimulation of the antigen-antibody complex and the pathogens, cascade reaction, which lead to activate complement C3. Then, membrane attact complex (MAC) is formed on the surface of the target cells, eventually crack the target cells. In addition, complement C3and its crack products have many functions of regulating immune system. Generally speaking, complement C3plays an important role in the natural immune system and the adaptive immune system.Lamprey is the representative of the jawless vertebrates, the evolutionary status is between the invertebrates and the vertebrates. Its unique evolutionary status and immune system are always attract the attention of immunologists. In recent years, variable lymphocyte recepters were found in lamprey which revealed a special adaptive immune system in lamprey. Complement C3was also found in lamprey.But the immune functions of complement C3in lampetra japonica is still not clear. In order to research the processes and characteristics of the immune system related complement C3of lampetra japonica, part of complement C3gene clone, protein expression and protein purification were worked, then preparation of the anti-C3polyclonal antibody. This laid solid foundation for the related funtions of complement C3of lampetra japonica.In this study, a part of complement C3coding region of lampetra japonica was cloned, the longth is918bp, then prokaryotic expression vector was made. After prokaryotic expression and recombinant protein purification, we got pure complement C3protein. After New Zealand rabbits immunity, antibody purification we obtained anti-C3polyclonal antibody. The ELISA and Western blotting assays were made. The result of ELISA showed that the valence of anti-C3antidy is1:160000. The result of Western blotting showed that the antidy can detect the nature C3in lamprey sera.In summary, we obtained anti-C3polyclonal antibody, and it provided a major tool for studying the immune system based on complement C3of lamprey.