Highly sensitive electrochemical immunosensor for golgi protein 73 based on proximity ligation assay and enzyme-powered recycling amplification

摘要

Golgi protein 73 (GP73) is a potential hepatocellular carcinoma serum marker with better sensitivity and specificity. In this study, a highly sensitive electrochemical immunosensor which combines the highly selective proximity ligation assay (PLA) and highly efficient enzyme-powered recycling amplification has been developed for GP73 determination. PLA is triggered by affinity binding of two labelled antibody-DNA (P1-RAb and P2-MAb) to target protein, resulting in increased specificity. The formed immunocomplex hybrids with DNA2 to realize the release of methylene blue labelled mononucleotides fragment with the help of exonuclease III, which acts as the power of recycling amplification. The fragment can diffuse easily to the nafion modified indium tin oxide electrode surface and a strong electrochemical signal can be detected. Under the optimized conditions, the enhanced electrochemical intensity has a linear relationship with the concentration of GP73 in the range of 0.3 pg mL(-1) to 6.0 ng mL(-1) with the detection limit of 0.10 pg mL(-1). The developed immunosensor has been applied to detect the GP73 in the clinical serum samples with satisfactory results. (C) 2018 Elsevier B.V. All rights reserved.