AccScience Publishing / MI / Online First / DOI: 10.36922/MI025130028
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C7Mab-2: A novel monoclonal antibody against mouse CCR7 established by immunization of the extracellular loop domain

Haruto Yamamoto1 ,  Hiroyuki Suzuki1* ,  Tomohiro Tanaka1 ,  Hiroyuki Satofuka1 ,  Mika K. Kaneko1 ,  Yukinari Kato1*
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1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, Sendai, Miyagi, Japan
MI, 025130028 https://doi.org/10.36922/MI025130028
Received: 29 March 2025 | Revised: 29 April 2025 | Accepted: 3 June 2025 | Published online: 2 July 2025
© 2025 by the Author(s). This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution 4.0 International License ( https://creativecommons.org/licenses/by/4.0/ )
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Abstract

The chemokine receptors possess seven transmembrane helices connected by an extracellular N‐terminal region, three extracellular loops (ECL1–3), three intracellular loops, and an intracellular C‐terminal region. Specific monoclonal antibodies (mAbs) against chemokine receptors for flow cytometry have been developed using Cell-Based Immunization and Screening, and the N-terminal peptide immunization methods. However, there are few reports on the establishment of anti-chemokine receptor mAbs through immunization with ECL peptides. Here, an anti-mouse C–C chemokine receptor type 7 (mCCR7) mAb, C7Mab-2 (rat immunoglobulin G2b, kappa), was established through immunization with the ECL3 peptide. C7Mab-2 demonstrated reactivity to mCCR7-overexpressed Chinese hamster ovary-K1 (CHO/mCCR7) cells in flow cytometry, which was inhibited by the ECL3 peptide. C7Mab-2 did not show cross-reactivity with other mouse CC, CXC, CX3C, and XC chemokine receptors. The dissociation constant value of C7Mab-2 was determined to be 2.8 × 10−9 M for CHO/mCCR7 cells. Furthermore, C7Mab-2 detected mCCR7 in immunohistochemistry. This strategy could accelerate the development of novel chemokine receptor mAbs with high affinity and specificity.

Keywords
Mouse C–C chemokine receptor type 7
Monoclonal antibody
Extracellular loop
Peptide immunization
Flow cytometry
Immunohistochemistry
Funding
This research was supported in part by the Japan Agency for Medical Research and Development (AMED) under Grant Numbers: JP24am0521010 (to Y. Kato), JP24ama121008 (to Y. Kato), JP24ama221339 (to Y. Kato), JP24bm1123027 (to Y. Kato), and JP24ck0106730 (to Y. Kato), and by the Japan Society for the Promotion of Science (JSPS) Grants-in-Aid for Scientific Research (KAKENHI) under Grant Numbers. 22K06995 (to H. Suzuki), 24K18268 (to T. Tanaka), 24K11652 (to H. Satofuka), and 25K10553 (to Y. Kato).
Conflict of interest
The authors declare that they have no competing interest.
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Microbes & Immunity, Electronic ISSN: 3029-2883 Print ISSN: 3041-0886, Published by AccScience Publishing
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