Construction, Expression, Purification and Biotin Labeling of a Single Recombinant Multi-Epitope Antigen for Double-Antigen Sandwich ELISA to Detect Hepatitis C Virus Antibody

Title: Construction, Expression, Purification and Biotin Labeling of a Single Recombinant Multi-Epitope Antigen for Double-Antigen Sandwich ELISA to Detect Hepatitis C Virus Antibody

Volume: 18 Issue: 8

Author(s): Jing He, Bingshui Xiu, Guohua Wang, Kun Chen, Xiaoyan Feng, Xiaoguo Song, Cuixia Zhu, Xiqin Yang, Guanzhong Bai, Shigan Ling and Heqiu Zhang

Affiliation:

Keywords: Antibody detection, biotin labeling, expression, hepatitis C virus, multi-epitope protein, purification, open reading frame, ELISA, Human Immunodeficiency Virus, BioSun software, epitopes, cloning, prokaryotic expression vector, antigenicity, ligation cycle, biotinylation, immunogenicities, multiple recombinant peptides, hydrophilicity, labeled antigen, C-terminal-modified recombinant proteinsAntibody detection, biotin labeling, expression, hepatitis C virus, multi-epitope protein, purification, open reading frame, ELISA, Human Immunodeficiency Virus, BioSun software, epitopes, cloning, prokaryotic expression vector, antigenicity, ligation cycle, biotinylation, immunogenicities, multiple recombinant peptides, hydrophilicity, labeled antigen, C-terminal-modified recombinant proteins

Abstract: Based on B cell epitope predictions, a recombinant antigen with multiple epitopes from four Hepatitis C Virus fragments (C, NS3, NS4 and NS5) were engineered. The recombinant gene was then highly expressed in E. coli. The nonmodified and C-terminal-modified recombinant proteins were used for coating and biotin labeling, respectively, to establish the double-antigen sandwich ELISA. Ten positive reference samples confirmed by the CHIRON RIBA HCV 3.0 SIA kit were detected positive, Forty one plasma samples were positive among samples from 441 volunteers, which indicated that the recombinant antigen could readily react well with plasma HCV antibody. As critical reagents of double-antigen sandwich ELISA, the recombinant multi-epitope antigen and the C-terminal-modified and biotin-conjugated antigen show good antigenicity. In this study, we provide a simple approach to produce multiple epitopes within one recombinant protein in order to avoid the costly expression of less-effective pools of multiple proteins, which is the conventional strategy of diagnostic antigen production for HCV antibody detection.

Cite this article as:

He Jing, Xiu Bingshui, Wang Guohua, Chen Kun, Feng Xiaoyan, Song Xiaoguo, Zhu Cuixia, Yang Xiqin, Bai Guanzhong, Ling Shigan and Zhang Heqiu, Construction, Expression, Purification and Biotin Labeling of a Single Recombinant Multi-Epitope Antigen for Double-Antigen Sandwich ELISA to Detect Hepatitis C Virus Antibody, Protein & Peptide Letters 2011; 18 (8) . https://dx.doi.org/10.2174/092986611795714014