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

被引：109

作者：

Chen, Jin ^{[1
,2
]}

Dalal, Ravindra V. ^{[3
]}

Petrov, Alexey N. ^{[2
]}

Tsai, Albert ^{[1
,2
]}

O'Leary, Sean E. ^{[2
]}

Chapin, Karen ^{[3
]}

Cheng, Janice ^{[3
]}

Ewan, Mark ^{[3
]}

Hsiung, Pei-Lin ^{[3
]}

Lundquist, Paul ^{[3
]}

Turner, Stephen W. ^{[3
]}

Hsu, David R. ^{[3
]}

Puglisi, Joseph D. ^{[2
]}

机构：

[1] Stanford Univ, Dept Appl Phys, Stanford, CA 94305 USA

[2] Stanford Univ, Sch Med, Dept Biol Struct, Stanford, CA 94305 USA

[3] Pacific Biosci Inc, Dept Res & Dev, Menlo Pk, CA 94025 USA

来源：

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA | 2014年 / 111卷 / 02期

基金：

美国国家卫生研究院;

关键词：

MODE WAVE-GUIDES; RIBOSOME; DYNAMICS; TRANSLATION;

D O I：

10.1073/pnas.1315735111

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

070301 [无机化学]; 070403 [天体物理学]; 070507 [自然资源与国土空间规划学]; 090105 [作物生产系统与生态工程];

摘要：

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.

引用

页码：664 / 669

页数：6

共 34 条

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