A novel anti-mouse CXCR1 monoclonal antibody, Cx1Mab-8, demonstrates nanomolar affinity in flow cytometry

CXC chemokine receptor 1 (CXCR1) is an important regulator for neutrophil granulocyte activation through binding to the ligand interleukin-8 (IL-8). Upon binding to IL-8, CXCR1 activates downstream signaling, critical for innate and adaptive immune responses. The IL-8-CXCR1 axis also plays an important role in tumor progression, especially in the tumor microenvironment. CXCR1 antagonists or anti-IL-8 monoclonal antibodies (mAbs) have been developed and evaluated in clinical trials for inflammatory diseases and tumors. In this study, we developed novel mAbs for mouse CXCR1 (mCXCR1) using the N-terminal peptide immunization. Among the established anti-mCXCR1 mAbs, Cx₁Mab-8 (rat IgG_2b, kappa) recognized mCXCR1-overexpressed Chinese hamster ovary-K1 (CHO/mCXCR1) and mCXCR1-overexpressed LN229 (LN229/mCXCR1) by flow cytometry. The dissociation constant (<i>K</i> _D) values of Cx₁Mab-8 for CHO/mCXCR1 and LN229/mCXCR1 were determined as 4.1 × 10^-10 M and 1.5 × 10^-9 M, respectively. These results indicated that Cx₁Mab-8 is useful for detecting mCXCR1 by flow cytometry with high affinity and could contribute to obtaining the proof of concept in preclinical studies.