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Proc Natl Acad Sci U S A. 2014 Jan 14;111(2):664-9. doi: 10.1073/pnas.1315735111. Epub 2013 Dec 30.

High-throughput platform for real-time monitoring of biological processes by multicolor single-molecule fluorescence.

Author information

1
Department of Applied Physics, Stanford University, Stanford, CA 94305.

Abstract

Zero-mode waveguides provide a powerful technology for studying single-molecule real-time dynamics of biological systems at physiological ligand concentrations. We customized a commercial zero-mode waveguide-based DNA sequencer for use as a versatile instrument for single-molecule fluorescence detection and showed that the system provides long fluorophore lifetimes with good signal to noise and low spectral cross-talk. We then used a ribosomal translation assay to show real-time fluidic delivery during data acquisition, showing it is possible to follow the conformation and composition of thousands of single biomolecules simultaneously through four spectral channels. This instrument allows high-throughput multiplexed dynamics of single-molecule biological processes over long timescales. The instrumentation presented here has broad applications to single-molecule studies of biological systems and is easily accessible to the biophysical community.

PMID:
24379388
PMCID:
PMC3896158
DOI:
10.1073/pnas.1315735111
[Indexed for MEDLINE]
Free PMC Article

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