Development of functional human monoclonal single-chain variable fragment antibody against HIV-1 from human cervical B cells
| Autor: | Gregg J. Silverman, John Rutherford, Marikka Elia, Carlos F. Barbas, Jody D. Berry, Rupert Kaul, Roberta P. Fuller, Sarah Gobuty, Francis A. Plummer |
|---|---|
| Rok vydání: | 2003 |
| Předmět: |
medicine.drug_class
Phagemid Immunology Molecular Sequence Data Immunoglobulin Variable Region Cervix Uteri HIV Antibodies Monoclonal antibody Flow cytometry law.invention law Genetics medicine Humans Amino Acid Sequence Gene Infectivity B-Lymphocytes biology medicine.diagnostic_test Base Sequence Antibodies Monoclonal Virology Molecular biology Kenya Recombinant Proteins Monoclonal biology.protein Recombinant DNA HIV-1 Leukocytes Mononuclear Female Antibody |
| Zdroj: | Hybridoma and hybridomics. 22(2) |
| ISSN: | 1536-8599 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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