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Affinity Binding-Induced Hg2+ Release and Quantum Dot Doping for General, Label-Free, and Homogenous Fluorescence Protein Assay
journal contributionposted on 2018-06-15, 00:00 authored by Yishen Tian, Chen Xin, Shuang Liu, Ying Liu, Shufeng Liu
Herein, a general protein conversion and analysis strategy was developed for homogeneous, label-free, and sensitive protein detection, on the basis of the affinity binding-induced Hg2+ release for protein conversion, and the succeeding Hg2+ doping-induced ZnSe quantum dot (QD) photoluminescence for signal readout. Two DNA motifs were designed, each of which was conjugated with a protein-specific recognition ligand. The mercury ions were initially introduced into one DNA motif by T-Hg2+-T interaction. The Hg2+ releasing was then accomplished after protein recognition-initiated strand exchange reaction between two DNA motifs. Then, the simultaneous incorporation of the released Hg2+ into ZnSe QD resulted in a doping-dependent fluorescence emission at 560 nm correlated with protein analysis. The protein assay is outperformed only by a simple one-step mixing operation but no separation or washing steps. Also, the use of doped QD as a fluorogenic reporter can avoid the fluorophore and/or quencher labeling, and eliminate complex DNA manipulation procedures for signal readout or amplification involved in most existing nucleic acid-based protein conversion and analysis methods. The versatile and sensitive detection toward multivalent proteins was verified with the detection limits achieved at 0.034 nM for anti-Dig antibody, 0.012 nM for streptavidin, and 0.025 nM for thrombin. Thus, it shows great promise for protein analysis to accommodate the applications in disease diagnosis, biomarker screening, and clinical medicine.
protein analysisDNA motifsacid-based protein conversionprotein conversiondoping-dependent fluorescence emissionHgprotein recognition-initiated strand exchange reactionDNA manipulation proceduresQuantum Dot DopingHomogenous Fluorescence Protein Assay Hereinsignal readoutprotein-specific recognition ligandQD