title: [Expression, purification and renaturation of Pol P51 antigen of HIV-1 strain CN54 and its application in antibody detection].
abstract: To obtain the pure and soluble P51 antigen of HIV-1 strain CN54, we transformed the Escherichia. coli strain BL21 codonplus-RIL with recombinant plasmid pTHioHisA51 which carries a gene encoding the Polymerase (Pol) P51 antigen of HIV-1 CN54 formerly, and induced protein expression by IPTG. We purified the recombinant protein with Chelating Sepharose FF-Ni and DEAE-Sepharose FF column chromatography, then renatured the recombinant protein by dialyzation. Purified protein was identified by Western blotting. We labeled and coated antigen P51 in a dual-antigen sandwich system, and tested it with serum samples from HIV-infected individuals. The results showed that P51 was expressed as inclusion body, and represented about 50% of total cellular protein. After purification and renaturation, the purity of P51 was up to 95%. Western blotting and sandwich ELISA demonstrated that recombinant P51 had good anti-HIV antibody specificity and sensitivity. The results suggested that recombinant HIV-1 P51 can be prepared as diagnostic reagent, and provides valuable support for HIV-1 detection and vaccine research.
authors.affiliation: College of Life Science and Biopharmaceutics, Shenyang Pharmaceutical University, Shenyang, China.
authors.forename: Jue
authors.initials: J
authors.lastname: Hou
authors.forename: Jing
authors.initials: J
authors.lastname: Sun
authors.forename: Zhiyong
authors.initials: Z
authors.lastname: Xu
authors.forename: Wenling
authors.initials: W
authors.lastname: Fan
authors.forename: Yixuan
authors.initials: Y
authors.lastname: Zhang
authors.forename: Yong
authors.initials: Y
authors.lastname: Liu
authors.forename: Yanling
authors.initials: Y
authors.lastname: Hao
mesh_terms: D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015483:HIV Antibodies / Q000097:blood* / Q000276:immunology; D015658:HIV Infections / Q000276:immunology* / Q000821:virology; D054303:HIV Reverse Transcriptase / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D015497:HIV-1 / Q000145:classification / Q000276:immunology*; D006801:Humans; D020673:Protein Renaturation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D012680:Sensitivity and Specificity
chemical_list: D015483:HIV Antibodies; D011994:Recombinant Proteins; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase