[Construction and screening of type 1 human immunodeficiency virus specific phage antibodies combinatorial library].
Human monoclonal antibodies to type 1 immunodeficiency virus (HIV-1) gp120 were generated from phage antibody combinatorial library.


METHODS
The human immunoglobulin heavy chain Fd and light chain k genes were amplified by half-nested PCR from PBMC of patient infected with HIV. Phage antibody combinatorial library was constructed with the Fd and k chain genes using Pcomb3 as vector. The affinity selection and ELISA were adopted for generating specific phage antibodies. Partial DNA of a positive clone was sequenced and its soluble Fab was expressed in E coli. HIV-1 specific phage antibodies combinatorial library were constructed using the Fd and k genes and Pcomb3 vector. The library capacity was about 1.95 x 10(7). The specific phage antibodies were highly enriched after three rounds of biopanning selection against HIV-1 gp120 and 32% positive clones were detected by ELISA screening. DNA fragment coding for CH1 and CL derived from a positive clone was sequenced and its product was successfully expressed as soluble Fab which was specific for HIV-1 gp120. The HIV-1 specific phage antibody combinatorial library, and human monoclonal antibodies to HIV-1 gp120 have been used as tools for screening of neutralizing antibody to HIV-1, and the methods seem to be very crucial and applicable.
He
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Research Center of Virology, Beijing Ditan Hospital, Beijing 100011.
Wang
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X
Liu
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S
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral / Q000276:immunology*; D001435:Bacteriophages / Q000276:immunology*; D001483:Base Sequence; D020650:Combinatorial Chemistry Techniques; D004279:DNA, Viral / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005803:Genes, Immunoglobulin / Q000276:immunology; D005822:Genetic Vectors; D015699:HIV Envelope Protein gp120 / Q000235:genetics* / Q000276:immunology; D015497:HIV-1 / Q000276:immunology*; D006801:Humans; D008969:Molecular Sequence Data; D019151:Peptide Library
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D004279:DNA, Viral; D015699:HIV Envelope Protein gp120; D019151:Peptide Library