Epitope Mapping of the Anti-Human CCR2 Monoclonal Antibody C 2 Mab-9.
Tomohiro TanakaGuanjie LiTeizo AsanoMika K KanekoHiroyuki SuzukiYukinari KatoPublished in: Monoclonal antibodies in immunodiagnosis and immunotherapy (2022)
CC chemokine receptor type-2 (CCR2) belongs to the G protein-coupled receptors superfamily, localized on cell surface of some immune-related cells, including monocytes and macrophages. CCR2 and its ligand CCL2 are involved in the progression of various diseases such as cancers. Therefore, CCR2-targeted monoclonal antibodies (mAbs) are needed for treatment and diagnosis. Previously, we successfully developed an anti-human CCR2 (hCCR2) mAb, C 2 Mab-9 (mouse IgG 1 , kappa), which is applicable for flow cytometry and immunocytochemistry. In this study, we investigated the critical epitope of C 2 Mab-9. We conducted enzyme-linked immunosorbent assay (ELISA) using several N-terminal peptides of hCCR2, and demonstrated that C 2 Mab-9 recognizes 11-29 and 21-39 amino acids of hCCR2. We further performed ELISA using 20 peptides, which include alanine substitution of hCCR2. C 2 Mab-9 lost the reaction to the alanine-substituted peptides of F23A, F24A, D25A, Y26A, and D27A. Among them, F23A, F24A, D25A, and Y26A did not block the C 2 Mab-9 reaction with U937 cells in flow cytometry. These results indicate that the critical binding epitope of C 2 Mab-9 includes Phe23, Phe24, Asp25, and Tyr26.
Keyphrases
- monoclonal antibody
- flow cytometry
- dendritic cells
- regulatory t cells
- induced apoptosis
- amino acid
- endothelial cells
- cell cycle arrest
- cell surface
- oxidative stress
- high throughput
- induced pluripotent stem cells
- pluripotent stem cells
- nuclear factor
- cell death
- endoplasmic reticulum stress
- binding protein
- inflammatory response
- transcription factor
- cell proliferation
- combination therapy
- molecular dynamics simulations