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Ultrahigh-resolution optical trap with single-fluorophore sensitivity.

ABSTRACT: We present a single-molecule instrument that combines a time-shared ultrahigh-resolution dual optical trap interlaced with a confocal fluorescence microscope. In a demonstration experiment, we observed individual single fluorophore-labeled DNA oligonucleotides to bind and unbind complementary DNA suspended between two trapped beads. Simultaneous with the single-fluorophore detection, we clearly observed coincident angstrom-scale changes in tether extension. Fluorescence readout allowed us to determine the duplex melting rate as a function of force. The new instrument will enable the simultaneous measurement of angstrom-scale mechanical motion of individual DNA-binding proteins (for example, single-base-pair stepping of DNA translocases) along with the detection of properties of fluorescently labeled protein (for example, internal configuration).

SUBMITTER: Comstock MJ

PROVIDER: S-EPMC3732480 | biostudies-literature | 2011 Apr

REPOSITORIES: biostudies-literature

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Ultrahigh-resolution optical trap with single-fluorophore sensitivity.

Comstock Matthew J MJ Ha Taekjip T Chemla Yann R YR

Nature methods 20110220 4

We present a single-molecule instrument that combines a time-shared ultrahigh-resolution dual optical trap interlaced with a confocal fluorescence microscope. In a demonstration experiment, we observed individual single fluorophore-labeled DNA oligonucleotides to bind and unbind complementary DNA suspended between two trapped beads. Simultaneous with the single-fluorophore detection, we clearly observed coincident angstrom-scale changes in tether extension. Fluorescence readout allowed us to det ...[more]

PMID: 21336286

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