Prussian blue nanoparticle-labeled aptasensing platform on graphene oxide for voltammetric detection of ɑ-fetoprotein in hepatocellular carcinoma with target recycling

An enzyme-free electrochemical aptasensing platform based on graphene oxide nanosheet-modified gold-disk electrode was developed for the voltammetric detection of alpha-fetoprotein (AFP) in hepatocellular carcinoma by using Prussian blue nanoparticle (PBNP)-labeled aptamer. The electroactive PBNP, a typical signal-generation tag, was utilized for the labeling of the aminated AFP aptamer by using the covalent conjugation. The electrochemical sensing platform was simply prepared on the basis of π-π stacking reaction between the immobilized graphene oxide and PBNP-labeled AFP aptamer. Upon target AFP introduction, the analyte reacted with the aptamer, thus resulting in the dissociation of the PBNP from the nanosheets. In the presence of DNase I, the newly formed AFP/aptamer-PBNP complex was cleaved to release target AFP, which could react again with the aptamer on the nanosheets, thereby causing the target recycling. During this process, the cleaved PBNP-aptamer was far away from the electrode to decrease the voltammetric signal. Under optimum conditions, the voltammetric peak current of the modified electrode decreased with the increment of target AFP concentration within the linear range of 0.01 – 300 ng mL-1 at a low detection limit of 6.3 pg mL-1. The precision and reproducibility of the aptasensing protocol were acceptable (CV: < 15% for intra-assay and inter-assay). Other possible nontarget biomarkers did not interfere significantly the voltammetric signal of this system. Human serum samples containing target AFP were assayed with the electrochemical aptsensing and commerical human AFP ELISA kit, and gave well-matched results from these two methods. Importantly, our strategy provides a new horizon for the determination of disease-related proteins.