title: [Prokaryotic expression and polyclonal antibody preparation of HPV6b E7 protein].
abstract: To express and prepare polyclonal antibody of Human papillomavirus type 6b (HPV6b) E7 protein. a prokaryotic expression vector pGEX-4T-2/HPV6b E7 was constructed and GST-HPV6b E7 fusion protein was expressed as a soluble protein in E. coli. The expressed fusion protein was purified via Glutathione-Sepharose 4B column and thrombin cleavage in order to obtain HPV6b E7 protein. Polyclonal IgG antibody was prepared by immunizing New-Zealand rabbits with HPV6b E7 protein. Western-Blot and immunofluorescence analysis showed that the polyclonal IgG antibody could specifically recognize HPV6b E7 protein and its titer was identified. SDS-PAGE analysis demonstrated that large amounts of soluble GST-HPV6b E7 fusion protein was expressed in E. coli after 3.0-6.0 hours of IPTG induction. Polyclonal IgG antibody successfully prepared from immunized rabbits showed high titer and high specificity as confirmed by Western-Blot and immunofluorescence. The preparation of anti-HPV6b E7 polyclonal antibody will facilitate further research on the biological and immunological functions of HPV6b E7 protein.
authors: Department of Dermatology and Venereology, School of Medicine, Sun Run Run Shaw Hospital, Zhejiang University, Hangzhou 310016, China. tangyi.hz@163.com
authors: Yi
authors: Y
authors: Tang
authors: Qiang
authors: Q
authors: Zhou
authors: Qi
authors: Q
authors: Wang
authors: Hao
authors: H
authors: Cheng
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology*; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D005260:Female; D015964:Gene Expression Regulation, Bacterial; D057809:HEK293 Cells; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology / Q000378:metabolism; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism
chemical_list: D000906:Antibodies; C085208:E7 protein, Human papillomavirus 6b; D007074:Immunoglobulin G; D009856:Oncogene Proteins, Viral; D011993:Recombinant Fusion Proteins