Molecular Encoded Beads with DNA Duplex Programming Fluorophore–Quencher Distance for Multiplexed Detection

Currently, the leading-edge bead encoding technique adopts the dye-doped strategy, which requires the construction of an encoded bead library in advance and is limited by strict material and technical barriers. Here, we report a simple, versatile, and plug-and-play bead encoding strategy, which uses bead surface DNA encoding instead of a dye-doped strategy. This is a fluorophore-quencher (F-Q) distance encoded strategy through the different Q-labeled positions at the DNA duplex, thereby changing the FRET efficiency. The strategy can simultaneously realize target capture and encoding on a bead surface, which greatly simplifies the process and reduces detection costs. In this work, we encoded 9-plex barcodes through the strategy and decoded them by 2-channel fluorescence flow cytometry. It is a modular technique that enables seamless integration with additional signal amplifier units. Herein, we constructed three kinds of multiplexed nucleic acid detection models, including "direct capture", "capture and then amplify", and "amplify and then capture", that enrich the encoding toolbox for bead-based multiplexed detection.