Development of electrochemical alpha-fetoprotein immunosensor

In this study, a new immunosensor was developed for the electrochemical determination of alpha-fetoprotein (AFP) that is a cancer biomarker and its application was performed on human blood serum samples. In the work, firstly the screen printed carbon electrodes (SPCE) were modified with reduced graphene oxide (RGO), then the electrode surface was coated with polyneutral red by applying the electropolymerization technique, and finally, gold nanoparticles (AuNP) were created by electrodeposition on the surface. Electrochemical characterizations of the SPCE/RGO/PNR/AuNP electrode were performed using cyclic voltammetry (CV), differential pulse voltammetry (DPV) and square wave voltammetry (SWV) methods. Immobilization of Anti-AFP to the surface of SPCE/RGO/PNR/AuNP electrodes were carried out by covalent binding method using 3-mercaptopropionic acid (3-MPA) and 1-ethyl-3-(3-dimethylamino-propyl) carbodiimide/N-hydroxy succinimide (EDC/NHS). The optimum working parameters of the prepared AFP immunosensor were determined by DPV and SWV methods. The optimum anti-AFP concentration, incubation time of Anti-AFP and incubation time of AFP were found to be 50.0 µg/mL, 60 min and 15 min, respectively. The analytical characterization of electrochemical AFP immunosensors were performed under the optimum working parameters. It was calculated that the linear ranges were 1-500 ng.mL-1, limit of detections (LOD) were 6.51 ng.mL-1 and 9.68 ng.mL-1, respectively. The reproducibility of the AFP immunosensor was tested for 50 ng.mL-1 AFP concentration and relative standard deviation (R.S.D.) values were calculated as 4.06% (n = 10) and 3.68% (n = 10) for DPV and SWV methods, respectively. The operation stability of the developed AFP immunosensor was examined by voltammetric methods for 60 days, and the storage stability for 12 weeks. The selectivity of the AFP immunosensor was tested by performing AFP analysis in the presence of 9 different antigen mixes and it was found that the antigen mixture made a 4.02% interference. The prepared AFP immunosensors were used for the analysis of different concentrations of AFP added to the human blood serum samples in a known amount and over 95% recoveries were obtained.