Ultrasensitive immunoassay of insulin based on highly efficient electrochemiluminescence quenching of carboxyl-functionalized g-C3N4 through coreactant dual-consumption by NiPd-DNAzyme

Abstract

An efficient quenching electrochemiluminescence (ECL) immunosensor for highly sensitive detection of insulin was constructed using carboxyl-functionalized g-C3N4 (C-g-C3N4) as an ECL emitter and hollow NiPd nanoparticles loading with G-quadruplex/hemin DNAzyme (NiPd-DNAzyme) as the dual-quenching probe. In this protocol, the as-prepared C-g-C3N4 served as both a luminophore and the matrix to immobilize capture antibody (Ab1) by amide linkage, which exhibiting strong ECL activity in the presence of the coreactant H2O2. The NiPd showed good peroxidase-activity and loading capacity for both G-quadruplex/hemin DNAzyme and detection antibody (Ab2). The dual-peroxidase nature of NiPd-DNAzyme composite highly promoted the reduction of H2O2, resulting in the consumption of the coreactant of C-g-C3N4 and obvious ECL quenching. Using insulin as a model analyte, the change of ECL intensity was logarithmically related to the concentration of the insulin in the range from 0.1 pg·mL−1 to 20.0 ng·mL−1 with a detection limit of 33 fg·mL−1. The wide detection range and high sensitivity resulted from the enhanced ECL emission and highly efficient quenching ability of NiPd-DNAzyme. Furthermore, the ECL immunosensor presented good stability, repeatability and selectivity, which demonstrate that it will be potential in clinical application.