Quantum-dot-encoded microbeads for multiplexed genetic detection of non-amplified DNA samples

摘要

Barcoding technologies have become the basis for a new generation of molecular diagnostic platforms for measuring biomarkers in a high-throughput, rapid, and sensitive manner. Thus far, researchers have mainly focused on preparing different types of barcodes but, in order to use them optimally in genomic-and proteomic-based applications, there is a need to understand the effect of barcode and assay parameters on their performance. Herein, quantum-dot barcodes are systematically characterized for the detection of non-amplified DNA sequences. The effect of capture probes, reporter probes, and target DNA sequence lengths are studied, as well as the effect of the amount of noncomplementary sequences on the hybridization kinetics and efficiency. From DNA denaturation to signal detection, quantum-dot-barcode assays require less than one hour to detect a target DNA sequence with a linear dynamic range of 0.02-100 fmol. Three optically distinct quantum-dot barcodes are used to demonstrate the multiplexing capability of these barcodes for genomic detection. These results suggest that quantum-dot barcodes are an excellent platform for multiplex, rapid, and sensitive genetic detection. A multiplex genetic assay is developed with the platform of quantum-dot-encoded microbeads for the detection of non-amplified, restriction-digested DNA samples. The parameters affecting the kinetics and efficiency of the sandwich hybridization assay are carefully examined, including the length of target fragments and probes, as well as the presence of noncomplementary sequences.