AccScience Publishing / MI / Online First / DOI: 10.36922/MI025220048
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Establishment of a novel anti-mouse CD73 monoclonal antibody C73Mab-9 by the Cell-Based Immunization and Screening method

Tomohiro Tanaka1 ,  Tomoko Sakata2,3 ,  Shiori Fujisawa1 ,  Haruto Yamamoto1 ,  Yu Kaneko1 ,  Mika K. Kaneko1 ,  Hiroyuki Suzuki1* ,  Yukinari Kato1,2*
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1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, 2-1, Seiryo-machi, Aoba-ku, Sendai, Miyagi, Japan
2 Department of PFI Antibody Drug Development, Tohoku University Graduate School of Medicine, 2-1, Seiryo-machi, Aoba-ku, Sendai, Miyagi, Japan
3 Pharma Foods International Co., Ltd., 1-49 Goryo-Ohara, Nishikyo-ku, Kyoto, Japan
MI, 025220048 https://doi.org/10.36922/MI025220048
Received: 11 June 2025 | Revised: 1 August 2025 | Accepted: 11 August 2025 | Published online: 2 September 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

Cluster of differentiation 73 (CD73), also known as ecto-5’-nucleotidase, is a glycosyl-phosphatidylinositol-anchored cell-surface molecule that belongs to the 5’-nucleosidase family. CD73 metabolizes extracellular adenosine (ADO) 5’-monophosphate into ADO and inorganic phosphate. CD73 plays a vital role in cancer-immune evasion by facilitating ADO production. Therefore, the development of specific anti-CD73 antibodies is desired to clarify the biological and pathological functions of CD73. Moreover, developing antibodies against mouse CD73 (mCD73) is also essential for pre-clinical experiments. This study successfully established an anti-mCD73 monoclonal antibody (clone C73Mab-9, rat IgG2a, and lambda) using the Cell-Based Immunization and Screening (CBIS) method. In flow cytometric analysis, C73Mab-9 was confirmed to recognize mCD73 in mCD73-overexpressed CHO-K1 (CHO/mCD73), MUSS, and NMuMG cells. The dissociation constant values of C73Mab-9 were 7.6 × 10−9 M for CHO/mCD73 and 1.2 × 10−9 M for MUSS cells, respectively. Furthermore, C73Mab-9 detected mCD73 in Western blot analysis. Therefore, C73Mab-9, established by the CBIS method, is useful for basic research and is expected to contribute to pre-clinical studies.

Keywords
Cluster of differentiation 73
Cell-based immunization and screening method
Monoclonal antibody
Flow cytometry
Funding
This research was supported in part by the Japan Agency for Medical Research and Development (AMED) under Grant Numbers: JP25am05210 (to Y. Kato), JP25ama121008 (to Y. Kato), JP25ama221339 (to Y. Kato), and JP25bm1123027 (to Y. Kato), and by the Japan Society for the Promotion of Science (JSPS) Grants-in-Aid for Scientific Research (KAKENHI) under Grant Numbers 24K18268 (to T. Tanaka) and 25K10553 (to Y. Kato).
Conflict of interest
The authors declare that they have no competing interests.
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