Copper nanocluster-labeled hybridization chain reaction for potentiometric immunoassay of matrix metalloproteinase-7 with acute kidney injury and renal cancer

A simple and portable potentiometric immunoassay was designed for the quantitative detection of matrix metalloproteinase-7 (MMP7) in acute kidney injury and renal cancer on copper ion-selective electrode (Cu-ISE). To construct such an assay system, the sandwich-type immunoreaction was initially carried out on the monoclonal anti-MMP7 capture antibody-coated microplates by using initiator DNA strand-labeled anti-MMP7 secondary antibody. Thereafter, a hybridization chain reaction (HCR) was used for the signal amplification between copper nanocluster-labeled two hairpin DNA probes. Accompanying formation of the sandwiched immunocomplex, the labeled DNA initiator strand on the secondary antibody triggered the HCR reaction between two alternating DNA hairpins to form a nicked double-helix with copper nanoclusters. Subsequent potentiometric measurement of copper ions released from the concatenated copper nanoclusters under acidic condition was conducted on a portable handheld potentiometer. The immuno-HCR assay combined easy-operation potentiometric assay with the signal amplification by using HCR-induced copper nanocluster concatamer. Under optimum conditions, the developed immuno-HCR assay exhibited a dynamic linear range of 0.01 – 100 ng mL-1 with a detection limit of 5.3 pg mL-1 and good reproducibility. Also, the immuno-HCR assay gave good anti-interferring capacity toward other biomarkers. The analysis for clinical human serum specimens revealed good accordance with the results obtained by an enzyme-linked immunosorbent assay. Importantly, our strategy is promising for enzyme-free and cost-effective analysis of low-aboundance cancer biomarkers.