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This work integrates target-induced DNA assembly and cleavage on a DNA chip to design a versatile imaging strategy as an assay for multiple proteins. The DNA assembly is achieved via immunological recognition to trigger the proximity hybridization for releasing a DNA sequence, which then hybridizes with FITC-DNA1 immobilized on the chip to induce the enzymatic cleavage of DNA1 and thus decrease the signals. The signal readout is performed with both fluorescent imaging of the left FITC and chemiluminescent (CL) imaging, by adding peroxidase labelled anti-FITC in assembly solution and CL substrates to produce CL emission. This one-step incubation can be completed in 30 min. The imaging method shows wide detection ranges and detection limits down to pg mL -1 for the simultaneous detection of 4 protein biomarkers. This high-throughput strategy with good practicability can be easily extended to other protein analytes, providing a powerful protocol for protein analysis and clinical diagnosis.

High-throughput imaging assay of multiple proteins via target-induced DNA assembly and cleavage