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Microbes & Immunity

COMMUNICATION
Ea Mab-9: A novel monoclonal antibody against
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erythropoietin-producing hepatocellular
receptor A8 for flow cytometry

Tomohiro Tanaka , Haruto Yamamoto , Yu Kaneko, Keisuke Shinoda,
Takuya Nakamura, Guanjie Li , Shiori Fujisawa, Hiroyuki Satofuka ,
Mika K. Kaneko , Hiroyuki Suzuki* , and Yukinari Kato*
Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, Sendai,
Miyagi, Japan

Abstract

Erythropoietin-producing hepatocellular receptor A8 (EphA8) is a type I
transmembrane protein that belongs to the largest erythropoietin-producing
hepatocellular (Eph) family among receptor tyrosine kinases. By binding to its
membrane-bound ephrin-A or ephrin-B ligands on adjacent cells, Eph receptors
form complexes and mediate bidirectional signaling activities, triggering cell-
cell adhesion and repulsion. Increased expression of EphA8 correlates with poor
*Corresponding authors: prognosis in some types of cancer. Therefore, developing sensitive monoclonal
Hiroyuki Suzuki
(hiroyuki.suzuki.b4@tohoku.ac.jp) antibodies (mAbs) for EphA8 has been desired for treatment, diagnosis, and further
Yukinari Kato basic research. In particular, there are no anti-EphA8 mAbs that can be used for flow
(yukinari.kato.e6@tohoku.ac.jp) cytometry. A novel, specific, and sensitive anti-human EphA8 mAb, which applies to
Citation: Tanaka T, Yamamoto H, flow cytometry, clone Ea Mab-9 (mouse immunoglobulin G , kappa), was established
1
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Kaneko Y, et al. Ea Mab-9: A novel using the Cell-Based Immunization and Screening method. Ea Mab-9 reacted with
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monoclonal antibody against 8
erythropoietin-producing EphA8-overexpressed Chinese hamster ovary-K1 cells (CHO/EphA8) and EphA8-
hepatocellular receptor A8 for flow overexpressed LN229 glioblastoma cells (LN229/EphA8) in flow cytometry. Notably,
cytometry. Microbes & Immunity. Ea Mab-9 did not recognize other members of the Eph receptor family. Furthermore,
2025;2(4):150-160. 8
doi: 10.36922/MI025060010 Ea Mab-9 demonstrated a high binding affinity for CHO/EphA8 and LN229/EphA8,
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with dissociation constants of 1.3 × 10 M and 1.6 × 10 M, respectively. The
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-9
Received: February 3, 2025
reaction of Ea Mab-9 with CHO/EphA8 was completely blocked by a recombinant
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1st revised: March 28, 2025 EphA8 protein. Ea Mab-9 could be useful for analyzing the EphA8-related biological
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2nd revised: May 20, 2025 responses using flow cytometry, owing to its high affinity and specificity.
Accepted: May 28, 2025
Keywords: EphA8; Cell-Based Immunization and Screening method; Monoclonal
Published online: June 18, 2025
antibody; Flow cytometry
Copyright: © 2025 Author(s).
This is an Open-Access article
distributed under the terms of the
Creative Commons Attribution
License, permitting distribution, 1. Introduction
and reproduction in any medium,
provided the original work is Erythropoietin-producing hepatocellular (Eph) receptors are the most prominent
properly cited. family of receptor tyrosine kinases (RTKs) and regulate tissue homeostasis, including
Publisher’s Note: AccScience cell proliferation and migration, tissue remodeling, angiogenesis, axon guidance, and
Publishing remains neutral with synaptic plasticity in the nervous system. Eph receptors are classified into EphA and
1-3
regard to jurisdictional claims in
published maps and institutional EphB subfamilies according to the sequence homology and binding mode to membrane-
affiliations. bound ephrin ligands. The EphA subfamily includes nine members, such as EphA1
Volume 2 Issue 4 (2025) 150 doi: 10.36922/MI025060010