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| Expression of Herpes Simplex Virus Type I Glycoprotein B in Eukaryotic Cells and Analysis of Its Antigenicity and Immunogenicity |
| WANG Zheng-mao1,LI Lin2,GUAN Wen-yan2,LI Yue-xi1,2 |
1.East-China Institute for Medical Biotechniques, Nanjing 210002,China
2.Department of Biochemistry & Molecular Biology,School of Preclinical Medicine, Nanjing Medical University, Nanjing 210029, China |
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Abstract To express and purify herpes simplex virus type I (HSV I) glycoprotein B (gB) in eukaryote cells with the purpose of analyzing the antigenicity and immunogenicity. At the beginning, the extracellular domain fragment gene of gD1 was synthesized by chemical method and cloned into eukaryotic expression vector pCEP4 to construct the recombinant plasmid pCEP4-gD1. After transfection of HEK293 cells with the recombinant plasmid, the expressed protein was characterized by Western blot and purified through Ni affinity chromatography. Then antigenicity of the protein was detected by ELISA. Finally, the purified protein was used to immunize Kunming mice in 1, 3, 5 weeks respectively, and antiserum were collected in 3, 5 and 7 weeks. The antibody titer was detected by indirect ELISA for immunogenicity analysis. Gene sequencing analysis demonstrates that the recombinant plasmid pCEP4-gB1 was constructed successfully. Western blot analysis indicated one major protein band, which molecular weight is approximate 85 kDa corresponding to the truncated forms of gB1 protein, was observed. In addition, ELISA detection showed that expressed gB1 has good antigenicity. After the third immunization, antibody titer of the mouse anti-gB1 was 5×103. The successful expression of the recombinant protein gD1, which can induce humoral immune response, lays a foundation for serological diagnosis and vaccine study of HSV.
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Received: 11 January 2010
Published: 12 June 2010
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