Development of an Anti-Mouse CCR8 Monoclonal Antibody (C8Mab-1) for Flow Cytometry and Immunocytochemistry.

Abstract

It has been widely accepted that monoclonal antibody (mAb) is an effective tool for cancer immunotherapy. The C-C motif chemokine receptor 8 (CCR8) is highly expressed in regulatory T cells and many cancers and is associated with the progression of the cancers. However, its role in cancer progression remains unclear. Thus, the development of mAbs for CCR8 leads to cancer immunotherapy and elucidation of unknown mechanisms of CCR8-dependent cancer progression. In this study, we have developed an anti-mouse CCR8 (mCCR8) mAb (clone C₈Mab-1, rat IgG_2a, kappa) using the Cell-Based Immunization and Screening (CBIS) method. We showed that C₈Mab-1 and its recombinant antibody (recC₈Mab-1) bind to mCCR8-overexpressed Chinese hamster ovary (CHO)-K1 cells (CHO/mCCR8), but not to the parental CHO-K1 cells, in flow cytometry and immunofluorescence. Moreover, C₈Mab-1 and recC₈Mab-1 specifically reacted to P388 (a mouse lymphocyte-like cells) and J774-1 (a mouse macrophage-like cells), which express endogenous mCCR8, in both applications. These results suggest that C₈Mab-1, developed using the CBIS method, is useful for flow cytometry and immunocytochemistry against exogenous and endogenous mCCR8.