Microbes and Infection
Volume 9, Issue 8, July 2007, Pages 1026-1033
Aizhi Zhao, Weijun Qin, Yueheng Han, Weihong Wen, Wenhong Zhang, Zhonghui Lian, Gang Chen, Zhuoli Zhang, Jianqiang Peng, He Wang, Yinglu Guo.
Abstract
To develop reagents for early diagnosis and therapeutic drugs against SARS-associated coronavirus (SARS-CoV), a large (3 × 109) immunized human antibody library was constructed from peripheral blood mononuclear cells from six SARS convalescent patients. A single chain variable fragment antibody (N18) with high affinity against N protein of SARS-CoV was isolated. Sequence analysis revealed that the VL gene was composed of VL3 h (V lambda subgroup) and JL2 regions and the VH gene was composed of VH1-69 (VH1 subgroup), D2-15, D3-22 and JH6 regions. Soluble N18 antibody was expressed in Escherichia coli HB2151, purified by Ni–NTA affinity chromatography and verified by SDS-PAGE and Western blot. The potential application for early diagnosis was evaluated using N protein capture ELISA in which N18 antibody demonstrated high sensitive activity in detecting N protein of SARS-CoV. Finally, the potential usefulness of the N18 antibody in prophylaxis, vaccine design and therapy of SARS is discussed.
Keywords
SARS-CoV, scFv, Nucleocapsid, Phage display