Development of an Immunosensor for Detection of Lung Cancer Biomakers

Abstract

Early detection of cancer (long before the tumor is perceived) greatly increases the chance of curability, and is the point of focus for many cancer researchers. The mortality rate due to lung cancer has not decreased for a number of years. This rate can be reduced by early diagnosis, for example if the serum proteins, microRNA, tumor-associated antigens, Carcinoembryonic antigen (CEA) and YES1 are detected in elevated amounts in the blood of potential lung cancer patients using a highly sensitive biosensor. In this study an innovative and ultrasensitive sandwich electrochemical multiplex immunosensor was fabricated for the simultaneous, sensitive, specific, and rapid detection of lung cancer biomarkers CEA and YES1. CEA is a paramount broad-spectrum tumor marker and YES1 (v-YES-1 Yamaguchi sarcoma viral oncogene homolog 1), a member of the SRC (proto-oncogene tyrosine-protein kinase Src) family kinases (SFKs), has been found in a significant subset of patients with lung cancer. The combination of the duplex tumor biomarkers detection was realized to be a formidable perfect tool for screening, diagnosis and monitoring of lung cancer. Gold Nano clusters coupled with thiolated protein G provided a good conductive platform with high surface area, and provided more binding sites for both YES1 and CEA antibodies. Glutaraldehyde facilitated the cross linking of the antibodies to the electrode surface through the thiolated protein G. The Horseradish peroxidase (HRP)-functionalized Au nanoparticles served as a good conductive platform to encapsulate a large amount of redox probe and to label secondary antibodies for signal amplification, due to the bountiful reactive oxygen functional groups on its surface. Through differential pulse voltammetric (DPV) measurements, two separate signals were detected directly in a single run, which indicated the presence of YES1 and CEA. Under optimized conditions, the electrochemical immunosensor manifested good sensitivity and selectivity for the simultaneous detection of YES1 and CEA with linear ranges of 0.1–50 ng/mL. The detection limit for YES1 and CEA was found to be 0.0022 and 0.0034 ng/mL respectively. To assess the functionalization of the immunosensor for the investigation of YES1 and CEA in human samples, investigation was done in experimentally cultured cancer cell supernatants, cellular protein lysates and human plasma. The results indicated that the immunosensor was able to pick out the CEA and YES1 biomarker, and that the results correlated satisfactorily with those of enzyme linked immunosorbent assay (ELISA). This method gave an auspicious simple, sensitive and quantitative approach for the detection of NSCLC biomarkers. We recommend that that the immunosensor be further tested in vivo in other human sample specimens, for example bronchial lavage, urine or sputum and also that the immunosensor be miniaturized in order to develop a point of care gadget that can be used for clinical monitoring of cancer progression.