In the present study, a triple signal amplification strategy was developed for ultrasensitive immunosensing of prostate-specific antigen (PSA) tumor marker. The proposed system was achieved by modification of glassy carbon electrode with graphene oxide/chitosan film and covalently attached of monoclonal PSA antibody and thionine as redox probe onto the modified electrode surface. Then, immunosensing was completed using sandwich-type immunoreaction of the PSA–antigen between anti-PSA immobilized on the graphene/chitosan interface and PSA–aptamer. For improve the sensitivity, polyamidoamine dendrimer-encapsulated gold nanoparticles (AuNPs–PAMAM) was used for covalent attachment of PSA–aptamer and HRP linked aptamer (Au–PAMA/aptamer–HRP) as electrochemical label in the sandwich format and electrocatalytic reduction of H2O2 in the presence of enzymatically oxidized thionine was measured. Under optimized condition, the obtained detection limit and linear concentration range were 10 fg ml−1(S/N=3) and 0.1 pg ml−1 to 90 ng ml−1 respectively, using differential pulse voltammetry as measuring technique. In addition, electrochemical impedance spectroscopy (EIS) was used as simple, rapid, low cost label free analytical technique for PSA measurement with detection limit of 5 pg ml−1 at concentration range up to 35 ng ml−1. Finally, the immunosensor is used to PSA detection in human serum and prostate tissue samples and the obtained result is well agreed with the values obtained by the standard ELISA method. The obtained results indicate the proposed immunosensor can be used for monitor the differences in PSA concentration in cancer tissue samples which holds great promise in clinical screening of cancer biomarkers.
