An Anti-Glypican-2 Monoclonal Antibody, G₂Mab-7, for Flow Cytometry, Western Blotting, and Immunohistochemistry

Glypican-2 (GPC2) is a heparan sulfate proteoglycan that plays essential roles in enhancing re-ceptor signaling by recruiting heparan sulfate-binding ligands. Overexpression of GPC2 is ob-served in pediatric cancers, including neuroblastoma and medulloblastoma, where it supports oncogenic signaling and tumor progression. High-affinity GPC2 binders have been developed as effective immunotherapies targeting tumors with GPC2 overexpression. In this study, we gener-ated new anti-GPC2 monoclonal antibodies using a flow cytometry–based high-throughput screening. Among the clones, G2Mab-7 (IgG1, κ) selectively bound GPC2-overexpressing Chinese hamster ovary cells (CHO/GPC2), with no reactivity toward parental CHO-K1 cells in flow cy-tometry. Notably, G2Mab-7 also recognized endogenous GPC2 in the human neuroblastoma cell line IMR-32. Specificity testing showed that G2Mab-7 reacted with CHO/GPC2 and did not crossreact with other GPCs overexpressed in CHO-K1. The dissociation constant (KD) for G2Mab-7 was 2.3 × 10⁻⁸ M for CHO/GPC2 and 4.2 × 10⁻⁹ M for IMR-32, indicating moderate binding affinity. Additionally, G2Mab-7 can be used in western blotting and immunohistochem-istry. Overall, these findings show that G2Mab-7 is a versatile tool for basic research and a promising candidate for antibody-based therapies.