Establishment of Novel Anti-Mouse CCR3 Monoclonal Antibodies (C <sub>3</sub> Mab-6 and C <sub>3</sub> Mab-7) by N-terminal Peptide Immunization
Teizo Asano, Hiroyuki Suzuki, Nohara Goto, Tomohiro Tanaka, Mika K. Kaneko, Yukinari Kato
Abstract
The CC chemokine receptor 3 (CCR3) is a member of the G protein-coupled receptor family that is highly expressed in eosinophils and basophils. CCR3 has been proposed as a therapeutic target for human immunodeficiency virus and allergy diagnosis. Therefore, in this study, we developed specific and sensitive monoclonal antibodies (mAbs) for mouse CCR3 (mCCR3), which are useful for flow cytometry by peptide immunization. The established anti-mCCR3 mAbs, C 3 Mab-6 (rat IgG 1 , kappa) and C 3 Mab-7 (rat IgG 1 , kappa), reacted with mCCR3-overexpressed Chinese hamster ovary-K1 (CHO/mCCR3), in addition to mCCR3-endogenously expressed cell lines, such as P388 (mouse lymphoid neoplasma) and J774-1 (mouse macrophage-like) through flow cytometry. Kinetic analyses using flow cytometry indicated that the dissociation constants ( K D s) of C 3 Mab-6 for CHO/mCCR3, P388, and J774-1 cells were 8.7 × 10 −9 M, 1.4 × 10 −7 M, and 1.7 × 10 −7 M, respectively, whereas the K D s of C 3 Mab-7 for these cell lines were 3.7 × 10 −9 M, 5.1 × 10 −7 M, and 3.1 × 10 −7 M, respectively. Results also indicated that C 3 Mab-6 and C 3 Mab-7 are useful for detecting cells expressing CCR3 through flow cytometry, thereby making them potentially beneficial for treating CCR3-expressing cells.