Bioorthogonal chemistry-based high-efficient quantum dots binding boosts the detection sensitivity of plasmon-enhanced fluorescence platform for immunoassay

Although plasmon-enhanced fluorescence (PEF) technology has been demonstrated its potential in protein detection for disease early diagnosis, its detection sensitivity still has a big gap compared to that of methods featured with single-molecule detection ability. Herein, inspired by the properties of catalyst-free bioorthogonal chemistry between 1,2,4,5-tetrazine (Tz) and trans-cyclooctene (TCO) being capable of improving the nanoparticle binding efficiency, we initially incorporate semiconductor quantum dots (QDs) into PEF-based suspension microarray platform as the signal reporter and evaluate its performance for biomarker prostate-specific antigen (PSA) detection by comparing with traditional fluorescence protein-based methods. In our assay, Tz-tagged QDs could be readily absorbed onto the sandwich immunocomplex composed of capture antibody, target PSA and TCO-labelled detection antibody for signal output. To improve the QDs binding efficiency, detection performance obtained from QDs-conjugated detection antibody as the reporter prepared by the same bioorthogonal chemistry reaction was also evaluated. For the evaluation of its potential in practical application, the performance including detection sensitivity and dynamic range of the proposed method for PSA assay in clinical sera samples using commercially-available PEF-based planar microarray as the control was discussed, which revealed that the nanoparticle-binding efficiency base on traditional SA/avidin method could be dramatically improved by the proposed method.