Expression,purification and analysis of HIV-1 core p24 antigenicity

Objective To express,purify and analyze antigenicity of HIV-1 core p24 antigen in prokaryotic system.Methods HIV-1 p24 gene amplified by PCR from plasmid(BH-10) was subcloned into vector pET-22b(+) after enzymatic digestion.The recombinant plasmid was then transformed to(E.coli) host,BL21(DE3),and highly expressed after IPTG induction.Double enzyme digestion was used to confirm the correct insert in the recombinant plasmid.SDS-PAGE,Western blot and ELISA were used to analyze purity and antigenicity.Results PCR product and external gene section from the recombinant plasmid pET 22b-p24 showed the same size of 690 bp equal to p24 gene sequences.An external expressed protein band of Mr 26 ×10~3 was obtained after purified protein SDS-PAGE electrophoresis.Western blot showed recombinant protein had specific reaction with HIV-1 positive sera and no response with normal(sera).Sensitivity and specificity of ELISA were 93.94%(62/66) and 93.33%(28/30) respectively.(Conclusion) The recombinant p24 antigen constructed and expressed in E.coli had good antigenicity and potential to develop HIV confirmation reagent.