Document Detail


Isolation of human antibodies against hepatitis E virus from phage display library by immobilized metal affinity chromatography.
MedLine Citation:
PMID:  18348408     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
OBJECTIVE: To isolate human antibodies against hepatitis E virus from phage display library by a new method of panning phage antibody library based on immobilized metal affinity chromatography (IMAC). METHODS: Phage antibody library was allowed to mix with hex-His tagged expressed HEV specific antigen, NE2, in solution for adequate binding before affinity resin for hex-His was added. The non-specific phage antibodies were removed by extensive washing and the specific bound phage antibodies could then be eluted to infect TG1 or repeat the binding process for subsequent rounds of purification. The specificity of the selected human antibodies were tested by antigen competitive ELISA, human sera blocking ELISA, scFv expression, and sequence analysis. RESULTS: His-NE2 specific recombinant phages were successfully enriched after panning procedure. Two individual phage clones, 126 and 138, showed 50% inhibition in NE2 antigen competition ELISA and obvious blocking effect by HEV positive serum in blocking ELISA. Soluble scFv of 126, 138 bound to NE2 specifically. CONCLUSION: Two specific human phage antibodies against hepatitis E virus (HEV) from phage display library were isolated by immobilized metal affinity chromatography. The immobilized metal affinity chromatography applied to phage antibody selection was a helpful supplement to the selection in solution.
Authors:
Ying-Wei Chen; Wen-Xin Luo; Ming-Qiao Wang; Jin Wang; Li-Feng Li; Quan Yuan; Jun Zhang; Ning-Shao Xia
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Biomedical and environmental sciences : BES     Volume:  20     ISSN:  0895-3988     ISO Abbreviation:  Biomed. Environ. Sci.     Publication Date:  2007 Dec 
Date Detail:
Created Date:  2008-03-19     Completed Date:  2008-06-03     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  8909524     Medline TA:  Biomed Environ Sci     Country:  United States    
Other Details:
Languages:  eng     Pagination:  488-94     Citation Subset:  IM    
Affiliation:
National Institute of Diagnostic and Vaccine Development for infectious diseases, The Key Laboratory of Ministry of Education for Cell Biology and Tumor Cell Engineering, School of Life Sciences, Xiamen University, Xiamen 361005, Fujian, China.
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MeSH Terms
Descriptor/Qualifier:
Amino Acid Sequence
Antibodies, Viral / chemistry,  genetics,  immunology,  isolation & purification*
Bacteriophages / genetics
Chromatography, Affinity / methods*
Enzyme-Linked Immunosorbent Assay
Hepatitis E virus / immunology*
Humans
Imidazoles / chemistry
Metals
Molecular Sequence Data
Chemical
Reg. No./Substance:
0/Antibodies, Viral; 0/Imidazoles; 0/Metals

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