Expression and Purification of Human CD252 Gene and Analysis of Immunogenicity of Recombinant Protein

Objective To optimize the expression of the fusion protein with the human CD252 extracellular segment,purify the interest protein and prepare the monoclonal antibody of human CD252. Construct a solid basis for further studying. Methods The pET32a- CD252 recombinant plasmid sequenced correctly was transformed into a kind of strain which express BL21 ( DE3). The recombinant protein was induced by IPTG and identified by SDS-PAGE. In order to offer better parameters,the positive clones expression condition was optimized by adjusting the time and temperature of the induction. Purifying the interest protein using inclusion body depuration and SDS-PAGE. The BAlB/c mouse was immuned by the purified recombination protein. The titer of polyclonal antibody was identified by ELISA. Results The positive stain that expressed the pET32a-CD252 was obtained successfully. After induced by IPTG,SDS-PAGE showed that the product of in- terest protein and the now protein existed in inclusion body of bacterial. SDS-PAGE showed that fusion protein expressed highly when induced with 1.0mmol/L, IPTG for 4 hours at the temperature of 37℃ . After purifying the interest protein with inclusion body, the purified interest pro- tein was obtained. The titer of polyclonal antibody was 1:3200 by indirect ELISA. Conclusion The CD252 protein can be expressed by pro- karyotic expression vector of human CD252 extracellular gene,and the good condition of fusion protein was obtained. Furthermore,the expres- sion protein posses excellent immunogenicity.