Isolation of monoclonal antibodies with predetermined conformational epitope specificity.
Journal Article (Journal Article)
Existing technologies allow isolating antigen-specific monoclonal antibodies (mAbs) from B cells. We devised a direct approach to isolate mAbs with predetermined conformational epitope specificity, using epitope mimetics (mimotopes) that reflect the three-dimensional structure of given antigen subdomains. We performed differential biopanning using bacteriophages encoding random peptide libraries and polyclonal antibodies (Abs) that had been affinity-purified with either native or denatured antigen. This strategy yielded conformational mimotopes. We then generated mimotope-fluorescent protein fusions, which were used as baits to isolate single memory B cells from rhesus monkeys (RMs). To amplify RM immunoglobulin variable regions, we developed RM-specific PCR primers and generated chimeric simian-human mAbs with predicted epitope specificity. We established proof-of-concept of our strategy by isolating mAbs targeting the conformational V3 loop crown of HIV Env; the new mAbs cross-neutralized viruses of different clades. The novel technology allows isolating mAbs from RMs or other hosts given experimental immunogens or infectious agents.
Full Text
Duke Authors
Cited Authors
- Sholukh, AM; Mukhtar, MM; Humbert, M; Essono, SS; Watkins, JD; Vyas, HK; Shanmuganathan, V; Hemashettar, G; Kahn, M; Hu, S-L; Montefiori, DC; Polonis, VR; Schur, PH; Ruprecht, RM
Published Date
- 2012
Published In
Volume / Issue
- 7 / 6
Start / End Page
- e38943 -
PubMed ID
- 22737224
Pubmed Central ID
- PMC3380854
Electronic International Standard Serial Number (EISSN)
- 1932-6203
Digital Object Identifier (DOI)
- 10.1371/journal.pone.0038943
Language
- eng
Conference Location
- United States