Herein, a label-free and ultrasensitive electrochemical aptasensor for the determination of lead(II) (Pb2+) was described. It was based on the application of a N,P dual-doped carbon dot–chitosan (N,P-CD–CS) composite as the signal molecule carrier and an aptamer (APT) as the specific binding probe for Pb2+ that were self-assembled on the surface of a gold electrode (GE). 6-Mercapto-1-hexanol (MCH) was used to block the nonspecific binding sites, and the electro-active molecule thionine (THi) was used as the signaling probe. The differential pulse voltammetry (DPV) response of THi at a rather low working potential of −0.17 V (vs. Ag/AgCl) was used to detect Pb2+. The electrochemical performances of the resulting modified electrode were investigated by cyclic voltammetry and electrochemical impedance spectroscopy. Under optimal experimental conditions, the modified electrode exhibited excellent DPV response depending on the concentration of Pb2+ in the 0.01 nM to 10 nM range. The limit of detection was 3.8 pM (at S/N = 3). The modified electrode displayed good reproducibility and excellent stability. It was successfully applied for the determination of Pb2+ in real water samples.
In this study, hemin-functionalized polyethyleneimine/silica nanoparticles (hemin/PEI/SiO2 NPs) served as an electrochemical label for ultrasensitive detection of prostate-specific antigen (PSA), a clinically important prostate cancer biomarker. A poly(thionine)/carbon nanotube (PTh/CNTs) multilayer film was fabricated on a glassy carbon electrode (GCE) via integrated electropolymerization and electrostatic layer-by-layer (LbL) assembly, forming a {PTh/CNTs}2/PTh modified GCE with abundant amino groups. PSA capture antibodies (Ab1) were then covalently immobilized on the modified electrode through glutaraldehyde (GA) crosslinking, leveraging the film's surface-exposed amino groups. Furthermore, hemin/PEI/SiO2 NPs were synthesized through amine-carboxyl interactions between carboxyl groups of hemin and amine groups of PEI/SiO2 NPs. The immunosensor assembly employed a sandwich-type electrochemical platform using PSA detection antibody (Ab2)-conjugated hemin/PEI/SiO2 NPs. The specific antibody-antigen binding enables efficient electron transfer across the nanoscale spacing (several nanometers) between hemin/PEI/SiO2 NPs and the {PTh/CNTs}2/PTh multilayer film. In this architecture, CNTs serve as excellent electron mediators, significantly facilitating electron transport. Moreover, the high-density immobilization of hemin on PEI/SiO2 NPs significantly amplifies the electrochemical redox currents of the multilayer film through proximity effects. This enhanced electrical signal enables highly sensitive detection of target protein antigen–antibody binding events. Consequently, by correlating the enhanced reduction peak current of the immunosensor with PSA concentration, highly sensitive detection of PSA at various concentrations can be achieved. Under optimized conditions, it exhibited a linear response to logarithmic PSA concentrations (0.01 pg mL−1 to 100 pg mL−1, R = 0.9963), with an ultralow detection limit of 3.0 fg mL−1 (S/N = 3). Furthermore, the immunosensor demonstrated outstanding analytical performance in human serum samples (96.0–103% recovery, n = 3), confirming its high accuracy for clinical applications.
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