Affiliation:
1. Department of Chemistry, Soonchunhyang University, Asan 31538, Republic of Korea
Abstract
An electrochemically reduced graphene oxide (ERGO) electrode-based electrochemical assay was developed for rapid, sensitive, and straightforward analysis of both activity and inhibition of the endonuclease EcoRV. The procedure uses a DNA substrate designed for EcoRV, featuring a double-stranded DNA (dsDNA) region labeled with methylene blue (MB) and a single-stranded DNA (ssDNA) region immobilized on the ERGO surface. The ERGO electrode, immobilized with the DNA substrate, was subsequently exposed to a sample containing EcoRV. Upon enzymatic hydrolysis, the cleaved dsDNA fragments were detached from the ERGO surface, leading to a decrease in the MB concentration near the electrode. This diminished the electron transfer efficiency for MB reduction, resulting in a decreased reduction current. This assay demonstrates excellent specificity and high sensitivity, with a limit of detection (LOD) of 9.5 × 10−3 U mL−1. Importantly, it can also measure EcoRV activity in the presence of aurintricarboxylic acid, a known inhibitor, highlighting its potential for drug discovery and clinical diagnostic applications.
Funder
National Research Foundation of Korea
Ministry of Science and ICT
Soonchunhyang University
Subject
Clinical Biochemistry,General Medicine,Analytical Chemistry,Biotechnology,Instrumentation,Biomedical Engineering,Engineering (miscellaneous)