Integrated bio-affinity nano-platform into a microfluidic immunosensor based on monoclonal bispecific trifunctional antibodies for the electrochemical determination of epithelial cancer biomarker

Background: The epithelial cell adhesion molecule (EpCAM) is a biomarker that is highly overexpressed on the surface of epithelial carcinoma cells. In this study, silver nanoparticles covered with polyvinyl alcohol (AgNPsPVA) were synthesized, characterized and used in a microfluidic immunosensor based on the use of anti-EpCAM recombinant antibodies as a trapping agent. Methods: The concentration of trapped EpCAM is then electrochemically quantified by HRP-conjugated anti-EpCAM-antibody. HRP reacted with its enzymatic substrate in a redox process which resulted in the appearance of a current whose magnitude (at a working voltage as low as 0.10 V) is directly proportional to the concentration of EpCAM. Results: Under optimized conditions, the detection limits for the microfluidic immunosensor and a commercial ELISA were 0.8 and 13.9 pg/L, respectively. The within-assay and between-assay coefficients of variation are below 6.5% for the proposed method. The immunosensor was validated by analyzing patient samples, and a good correlation with a commercial ELISA was obtained. Conclusions: The good analytical performance is attributed to the efficient immobilization of the anti-EpCAM recombinant antibodies on the AgNPs-PVA, and its high specificity for EpCAM. This microfluidic immunosensor is intended for use in diagnosis and prognosis of epithelial cancer, to monitor the disease, and to assess therapeutic efficacy. (C) 2016 Elsevier B.V. All rights reserved.