Fab’ Fragment-Immobilized Gold Surface for Capturing EpCAM-Positive Breast Cancer Cells

Abstract

ABSTRACT: Circulating tumor cells (CTCs) are cancer cells present in the bloodstream that originate from primary or metastatic sites. Sensitive and selective capture of these rare cells is essential for early diagnosis, metastasis prevention, and prognosis prediction. In this study, we demonstrated the effectiveness of a surface functionalized with epithelial cell adhesion molecule (EpCAM) Fab’ (fragment-antigen-binding) fragments for the specific capture of EpCAM-positive human breast cancer cells. EpCAM antibody Fab’ fragments were produced through pepsin digestion and characterized by SDS-PAGE analysis. Glass surfaces were silanized before being coated with a thin layer of gold via sputtering to ensure stability. The Fab’ fragments were immobilized on the gold-coated glass surfaces through strong gold-thiol bonds. The modified surfaces were then characterized using Fourier-transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and atomic force microscopy (AFM) analyses. Cell capture performance was assessed using fluorescence microscopy with both EpCAM-positive and EpCAM-negative cell lines. The results show that the Fab’-modified surface offers a promising platform for the selective immunocapture of EpCAM-positive cells. Practical application: This study presents a preliminary design of a Fab’ fragment-immobilized surface for the selective capture of EpCAM-positive breast cancer cells. The surface modification relies on spontaneous Au-S bonding, offering a simple and effective chemical method. The modified surface demonstrates strong potential for integration into future biosensor platforms for detecting circulating tumor cells. Such a system is promising for advanced diagnostics, monitoring, disease progression, and personalized treatment uses. © 2025 Elsevier B.V., All rights reserved.