High-Fidelity Single Molecule Quantification in a Flow Cytometer Using Multiparametric Optical Analysis.

ABSTRACT: Microfluidic techniques are widely used for high-throughput quantification and discrete analysis of micron-scale objects but are difficult to apply to molecular-scale targets. Instead, single-molecule methods primarily rely on low-throughput microscopic imaging of immobilized molecules. Here we report that commercial-grade flow cytometers can detect single nucleic acid targets following enzymatic extension and dense labeling with multiple distinct fluorophores. We focus on microRNAs, short nucleic acids that can be extended by rolling circle amplification (RCA). We labeled RCA-extended microRNAs with multicolor fluorophores to generate repetitive nucleic acid products with submicron sizes and tunable multispectral profiles. By cross-correlating the multiparametric optical features, signal-

SUBMITTER: Smith LD 

PROVIDER: S-EPMC7295608 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature