Goose Iga Heavy Chain Costont Region Prokaryotic Expression And Preparation And Identiifcation Of IgA Specific Antibodies

Immunoglobulin (Ig) is an important effector molecule of humoral immunity, which plays animportant role in immune regulation. There has a little report of the goose immunoglobulin so far atmolecule level.According to goose IgA heavy chain constant region (F) gene sequences, a pair of primers wasdesigned. PCR was carried to clone F genes and the prokaryotic expression vector pET-28-GoIgA-Cabout1339bp was constructed. The recombinant protein F was expressed in E. coliRosettaTM(DE3)plysS and the PAb against F was prepared with protein F.The antigen-antibody reactionbetween the PAb against F and others confirmed the success of protein F.It was laid the materialfoundation in order to further study the goose IgA structure and functional characteristics.The BALB/c mice were immuned with goose serum Ig purfied by Protein A affinitychromatography system. After three times of immunity, spleen cells of BALB/c mice which are collectedand SP2/0were fused. By I-ELISA, four hybridoma cell lines (3A4,2F12,4D4and4F12) were selectedto secret antibody against goose Ig. Four MAbs were blonging to IgG isotype,in which three McAbswere blonging to IgG1isotype, one McAbs were blonging to IgG2b isotype. Four MAbs have ability ofproducing antibodies against goose Ig in vitro. Then the biological characterization was identified. Theresults of indirect immunofluorescence, Western blot and indirect ELISA showed that the3A4and2F12hybridoma cell lines secret antibody against the recombinant protein F with high specificity, having noreaction with the Ig of other species.In summary, we cloned the gene of F and expressed recombinant protein containing F for the firsttime.Then the polyclonal antibody against the recombinant protein F was prepared for the first time,subsequently it was tested with specific identificationthe. The four MAbs were identified by using theobtained F recombinant protein. It showed that two MAbs were against the goose IgA heavy chainconstant region.Which laid the foundation for the research of goose basic immunology.