Development of Novel Anti-CDH1/E-Cadherin Monoclonal Antibodies for Flow Cytometry, Western Blotting, and Immunohistochemistry

Cadherin (CDH)-mediated extracellular homophilic binding is crucial for maintaining tissue homeostasis. The epithelial cell-cell adhesion molecule cadherin 1 (CDH1/E-cadherin) forms the adherens junctions in epithelial cells, and the loss of CDH1 facilitates the migration and invasion of carcinoma cells. Although several anti-CDH1 monoclonal antibodies (mAbs) are available for western blotting and immunohistochemistry (IHC), a highly sensitive anti-CDH1 mAb suitable for flow cytometry has not been developed. We developed anti-CDH1 monoclonal antibodies through a flow cytometry-based high-throughput screening. Two anti-CDH1 mAb clones, Ca1Mab-3 (IgG1, κ) and Ca1Mab-5 (IgG1, κ), reacted with human CDH1-overexpressed Chinese hamster ovary-K1 (CHO/CDH1) cells in flow cytometry. Furthermore, Ca1Mab-3 and Ca1Mab-5 recognize endogenous CDH1-expressing human luminal-type breast cancer cells, such as MCF-7, but not triple-negative breast cancer cells, like MDA-MB-231. The dissociation constant values of Ca1Mab-3 and Ca1Mab-5 for CHO/CDH1 were determined as 4.8 × 10−10 M and 1.3 × 10−9 M, respectively. Ca1Mab-3 and Ca1Mab-5 can detect endogenous CDH1 in western blotting and IHC using a cell block. Furthermore, Ca1Mab-5 is available for IHC in formalin-fixed paraffin-embedded tumor tissues. These results indicate that Ca1Mab-3 and Ca1Mab-5, established by the CBIS method, are versatile for basic research and are expected to contribute to clinical applications, such as tumor diagnosis and therapy.