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


  • eng

Conference Location

  • United States