Electrochemical cell-SELEX monitoring and its application to electrochemical aptasensor for colorectal cancer detection

Abstract

The cell systematic evolution of ligands by exponential enrichment (cell-SELEX) enabled the development of aptamers to recognize cell surface proteins in their embedded state for diagnostics and therapeutics. Traditional cell-SELEX monitoring relied on fluorescence-activated cell sorting (FACS) that requires expensive equipment. To overcome these limitations, this study proposed a simple and cost-effective, electrochemical cell-SELEX monitoring method. This method converted the binding reaction between cells and aptamers on the electrode into an electrical signal and analyzes it using electrochemical impedance spectroscopy (EIS). This method significantly reduced experimental time and costs by eliminating the need for expensive equipment and labeling with specific markers such as fluorescent dyes or enzymes. An aptamer selected through electrochemical cell-SELEX monitoring using colon cancer cell line SNU-81 shows high affinity (K_d of 101.5  nM). Affinity-based purification confirmed that the aptamer binds to heat shock protein 60 (HSP60). The affinity for HSP60 was further validated (K_d = 109.2 nM), and the similarity to the binding affinity observed with cells allowed for the identification of the aptamer’s binding site, demonstrating its potential as a biomarker for colorectal cancer (CRC). The developed colorectal cancer cell detection biosensor demonstrated a limit of detection of 88.7 cell/mL for concentrations within a range of 10² ––10⁶ cell/mL. Overall, this study establishes a streamlined cell-SELEX method widely applicable to biosensors and therapeutics. It provides a comprehensive development framework for cancer diagnostic devices, including aptamer development, biomarker discovery, and diagnostic sensor fabrication for colorectal cancer.