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

Jin Chen, Ravindra V. Dalal, Alexey N. Petrov, Albert Tsai, Seán E. O'Leary, Karen Chapin, Janice Cheng, Mark Ewan, Pei Lin Hsiung, Paul Lundquist, Stephen W. Turner, David R. Hsu, Joseph D. Puglisi

Research output: Contribution to journalArticlepeer-review

67 Scopus citations

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.

Original languageEnglish (US)
Pages (from-to)664-669
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume111
Issue number2
DOIs
StatePublished - 2014
Externally publishedYes

ASJC Scopus subject areas

  • General

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