Abstract
Since glucose biosensors are one of the most popular and widely used point-of-care testing devices, a novel electrochemical enzyme-linked immunosorbent assay (ELISA) for protein biomarkers has been developed based on a glucose detection strategy. In this study, α-fetoprotein (AFP) was used as the target protein. An electrochemical ELISA system was constructed using anti-AFP antibodies immobilized on microwell plates as the capture antibody (Ab1) and multi-label bioconjugates as signal tracer. The bioconjugates were synthesized by attaching glucoamylase and the secondary anti-AFP antibodies (Ab2) to gold nanoparticles (AuNPs). After formation of the sandwich complex, the Ab2-glucoamylase-AuNPs conjugates converted starch into glucose in the presence of AFP. The concentration of AFP can be calculated based on the linear relation between AFP and glucose, the concentration of which can be detected by the glucose biosensor. When the AFP concentration ranged from 0.05 to 100 ng/mL, a linear calibration plot (i (µA) = 13.62033 − 2.86252 logCAFP (ng/mL), r = 0.99886) with a detection limit of 0.02 ng/mL was obtained under optimal conditions. The electrochemical ELISA developed in this work shows acceptable stability and reproducibility, and the assay for AFP spiked in human serum also shows good recovery (97.0%–104%). This new method could be applied for detecting any protein biomarker with the corresponding antibodies.
Highlights
Detection of protein biomarkers for cancer and other fatal human diseases plays a key role in aiding an early diagnosis and monitoring disease progression [1,2,3]
The results suggested that the AFP concentration could be determined with the glucose-detection-based electrochemical enzyme-linked immunosorbent assay (ELISA) method
These results demonstrated the success of a novel electrochemical immunoassay based on glucoamylase mediator
Summary
Detection of protein biomarkers for cancer and other fatal human diseases plays a key role in aiding an early diagnosis and monitoring disease progression [1,2,3]. Perhaps the most successful example of such a device is the glucose meter, which is currently one of the most widely used diagnostic devices in the world, as a result of the more than 30 years of development [11,12,13] This widely used biosensor can only detect a single target, blood glucose. Future developments using glucose biosensors for protein biomarkers detection can potentially revolutionize the practice of medicine, with the ability to dramatically reduce healthcare costs. In this aspect, glucose biosensors could serve as general detectors used for both glucose and protein targets
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