Bioconjugation of peroxidase-like nanostructures with natural enzyme for in-situ amplified conductometric immunoassay of tissue polypeptide antigen in biological fluids

• We design a new conductometric immunoassay of tissue polypeptide antigen.
• Prussian blue-gold hybrid nanostructures with peroxidase-like activity were used as the labels.
• Natural enzyme and peroxidase-like mimics were utilized for signal amplification.
• Nano labels and enzyme labels were aggregated together.

A new conductometric immunoassay based on biofunctionalized prussian blue-gold hybrid nanostructure (PBGN) was designed for sensitive determination of tissue polypeptide antigen (TPA). The immunosensor was prepared by immobilizing anti-TPA capture antibody on the interdigitated transducer, while the PBGN was used for bioconjugation of detection antibody and horseradish peroxidase (HRP). Upon target TPA introduction, the sandwiched immunocomplex was formed between capture antibody and detection antibody on the transducer. Accompanying the PBGN, the carried HRP and prussian blue catalyzed iodine ion into iodine relative to H2O2-KI system, thus resulting in the variations of local conductivity. Under optimal conditions, the immunosensor exhibited good conductometric responses toward target TPA, and allowed the detectable concentration as low as 0.28 pg mL−1. The precision, specificity and reproducibility were acceptable. Importantly, our strategy was also utilized for analysis of 13 human serum specimens, giving well matched results with those obtained from human TPA ELISA kit.