Epitope mapping of an anti-human epidermal growth factor receptor monoclonal antibody (EMab-51) using the RIEDL insertion for epitope mapping method

Ren Nanamiya, Masato Sano, Teizo Asano, Miyuki Yanaka, Takuro Nakamura, Masaki Saito, Tomohiro Tanaka, Hideki Hosono, Nami Tateyama, Mika K. Kaneko, Yukinari Kato

Research output: Contribution to journalArticlepeer-review

Abstract

The classic method for identifying the epitope that monoclonal antibodies (mAbs) bind uses deletion mutants and point mutants of the target protein. However, determining the epitope of mAbs-reactive membrane proteins is often challenging. We recently developed the RIEDL insertion for epitope mapping (REMAP) method to identify mAb-binding epitopes. Herein, we first checked the reactivity of an anti-epidermal growth factor receptor (EGFR) mAb (EMab-51) to several EGFR deletion mutants such as EGFR/dN152, EGFR/dN313, EGFR/dN370, EGFR/dN375, EGFR/dN380, and EGFR/dN482. We found the N-terminus of the EMab-51-binding epitope between residues 375 and 380 of EGFR. We next produced EGFR/dN313 mutants with the RIEDL peptide tag inserted at each possible position of 375-AFRGDSFTHTPPLDP-389. EMab-51 lost its reactivity with the mutants having a RIEDL tag inserted at each position of 377-RGDSFTHTPP-386, whereas LpMab-7 (an anti-RIEDL mAb) detected every mutant. Thus, using the REMAP method, we identified the EMab-51-binding epitope of EGFR as 377-RGDSFTHTPP-386.

Original languageEnglish
Pages (from-to)149-155
Number of pages7
JournalMonoclonal antibodies in immunodiagnosis and immunotherapy
Volume40
Issue number4
DOIs
Publication statusPublished - 2021 Aug 1
Externally publishedYes

Keywords

  • EGFR
  • EMab-51
  • RIEDL tag
  • epitope mapping
  • monoclonal antibody

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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