Development of functional human monoclonal single-chain variable fragment antibody against HIV-1 from human cervical B cells

• Published in: Hybridoma and hybridomics Vol. 22; no. 2; p. 97
• Main Authors: Berry, Jody D, Rutherford, John, Silverman, Gregg J, Kaul, Rupert, Elia, Marikka, Gobuty, Sarah, Fuller, Roberta, Plummer, Francis A, Barbas, 3rd, Carlos F
• Format: Journal Article
• Language: English
• Published: United States 01.04.2003
• Subjects: Amino Acid Sequence; Antibodies, Monoclonal - genetics; Antibodies, Monoclonal - immunology; B-Lymphocytes - immunology; Base Sequence; Cervix Uteri - cytology; Cervix Uteri - immunology; Female; HIV Antibodies - genetics; HIV Antibodies - immunology; HIV-1 - immunology; Humans; Immunoglobulin Variable Region - genetics; Immunoglobulin Variable Region - immunology; Kenya; Leukocytes, Mononuclear - immunology; Molecular Sequence Data; Recombinant Proteins - genetics; Recombinant Proteins - immunology; Kenya
• ISSN: 1536-8599
• DOI: 10.1089/153685903321948021

A panel of novel recombinant single-chain variable fragment (scFv) antibody against human immunodeficiency virus type-1 (HIV-1) was isolated and characterized. We generated human scFvs using RNA harvested from cervical B lymphocytes of Kenyan prostitutes who are highly exposed to HIV-1, but remain persistently seronegative. The variable regions of the heavy (VH) and light (VL) chain antibody genes were selected as hybrids using guided-selection with the VL and VH, respectively, of a derivative of IgGb(12) using the phagemid vector pComb3X. IgGb(12) is a previously well-characterized HIV-1 neutralizing human monoclonal antibody (MAb). One of the hybrid scFv, IgA6/4L, neutralizes HIV-1 infectivity in in vitro cell culture assay. The cervical VH and VL chain antibody genes were connected by a DNA linker and subcloned in pComb3X. The cervical scFv clones were functional in recognizing HIV-1 gp120 by enzyme-linked immunosorbant assay (ELISA) and on cells in flow cytometry. Whole IgGb(12) does not inhibit binding of clones IgA6/5k nor IgA6/30lambda to gp120, which suggests that they bind different epitopes. Nucleotide sequence analysis of the cervical scFv show the clones are unique and reveal interesting characteristics of human cervical V gene pools. This work demonstrates, for the first time, cloning of a functional scFv MAb to a sexually transmitted disease pathogen from local cervical B-cell pools in exposed humans.