Single-molecule fluorescence resonance energy transfer in nanopipets: Improving distance resolution and concentration range

Vogelsang J, Doose S, Sauer M, Tinnefeld P (2007)
ANALYTICAL CHEMISTRY 79(19): 7367-7375.

Journal Article | Published | English

No fulltext has been uploaded

Author
; ; ;
Abstract
In recent years fluorescence resonance energy transfer (FRET) has widely been used to measure distances, binding, and distance dynamics at the single-molecule (sm) level. Some basic constraints of smFRET are the limited distance resolution owing to low photon statistics and the restriction to high affinity interactions. We demonstrate that by confining molecules in nanopipets with an inner diameter of similar to 100 nm at the tip, FRET can be measured with improved photon statistics and at up to 50-fold higher concentrations. The flow of the donor/acceptor (Cy3B/ATTO647N) labeled double-stranded DNA conjugates was established by electrokinetic forces. Because of the small inner diameter of the nanopipet, every molecule passing the tip is detected applying alternating laser excitation (ALEX). Thus, the technique offers the advantage to study interactions with smaller association constants (< 10(9) M-1) using minute sample amounts (< 5 mu L). The improved photon statistics reduces shot-noise contributions and results in sharper FRET distributions. Experimental results are supported by Monte Carlo simulations which also explain the occurrence of two populations in burst size distributions measured in nanopipet experiments. Because of the confinement of the molecules in nanopipets, the widths of FRET histograms are reduced to a degree where shot-noise is not the only limiting factor but also conformational dynamics of the linkers used to attach the chromophores have to be considered. In addition, our experiments emphasize the influence of photoinduced dark states on both the mean energy transfer efficiency and the width of FRET histograms.
Publishing Year
ISSN
eISSN
PUB-ID

Cite this

Vogelsang J, Doose S, Sauer M, Tinnefeld P. Single-molecule fluorescence resonance energy transfer in nanopipets: Improving distance resolution and concentration range. ANALYTICAL CHEMISTRY. 2007;79(19):7367-7375.
Vogelsang, J., Doose, S., Sauer, M., & Tinnefeld, P. (2007). Single-molecule fluorescence resonance energy transfer in nanopipets: Improving distance resolution and concentration range. ANALYTICAL CHEMISTRY, 79(19), 7367-7375.
Vogelsang, J., Doose, S., Sauer, M., and Tinnefeld, P. (2007). Single-molecule fluorescence resonance energy transfer in nanopipets: Improving distance resolution and concentration range. ANALYTICAL CHEMISTRY 79, 7367-7375.
Vogelsang, J., et al., 2007. Single-molecule fluorescence resonance energy transfer in nanopipets: Improving distance resolution and concentration range. ANALYTICAL CHEMISTRY, 79(19), p 7367-7375.
J. Vogelsang, et al., “Single-molecule fluorescence resonance energy transfer in nanopipets: Improving distance resolution and concentration range”, ANALYTICAL CHEMISTRY, vol. 79, 2007, pp. 7367-7375.
Vogelsang, J., Doose, S., Sauer, M., Tinnefeld, P.: Single-molecule fluorescence resonance energy transfer in nanopipets: Improving distance resolution and concentration range. ANALYTICAL CHEMISTRY. 79, 7367-7375 (2007).
Vogelsang, Jan, Doose, Soren, Sauer, Markus, and Tinnefeld, Philip. “Single-molecule fluorescence resonance energy transfer in nanopipets: Improving distance resolution and concentration range”. ANALYTICAL CHEMISTRY 79.19 (2007): 7367-7375.
This data publication is cited in the following publications:
This publication cites the following data publications:

15 Citations in Europe PMC

Data provided by Europe PubMed Central.

Breaking the concentration limit of optical single-molecule detection.
Holzmeister P, Acuna GP, Grohmann D, Tinnefeld P., Chem Soc Rev 43(4), 2014
PMID: 24019005
Single-molecule measurements of transient biomolecular complexes through microfluidic dilution.
Horrocks MH, Rajah L, Jonsson P, Kjaergaard M, Vendruscolo M, Knowles TP, Klenerman D., Anal. Chem. 85(14), 2013
PMID: 23782428
Role of conformational dynamics in α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor partial agonism.
Ramaswamy S, Cooper D, Poddar N, MacLean DM, Rambhadran A, Taylor JN, Uhm H, Landes CF, Jayaraman V., J. Biol. Chem. 287(52), 2012
PMID: 23115239
Nanochannel-based single molecule recycling.
Lesoine JF, Venkataraman PA, Maloney PC, Dumont ME, Novotny L., Nano Lett. 12(6), 2012
PMID: 22662745
Mechanisms and advancement of antifading agents for fluorescence microscopy and single-molecule spectroscopy.
Cordes T, Maiser A, Steinhauer C, Schermelleh L, Tinnefeld P., Phys Chem Chem Phys 13(14), 2011
PMID: 21311807
Single-molecule FRET ruler based on rigid DNA origami blocks.
Stein IH, Schuller V, Bohm P, Tinnefeld P, Liedl T., Chemphyschem 12(3), 2011
PMID: 21308944
Correlated movement and bending of nucleic acid structures visualized by multicolor single-molecule spectroscopy.
Person B, Stein IH, Steinhauer C, Vogelsang J, Tinnefeld P., Chemphyschem 10(9-10), 2009
PMID: 19499555
Photophysical aspects of molecular probes near nanostructured gold surfaces.
Ghosh SK, Pal T., Phys Chem Chem Phys 11(20), 2009
PMID: 19440609
Single-molecule photophysics of oxazines on DNA and its application in a FRET switch.
Vogelsang J, Cordes T, Tinnefeld P., Photochem. Photobiol. Sci. 8(4), 2009
PMID: 19337662
Nonequilibrium single molecule protein folding in a coaxial mixer.
Hamadani KM, Weiss S., Biophys. J. 95(1), 2008
PMID: 18339751

Export

0 Marked Publications

Open Data PUB

Web of Science

View record in Web of Science®

Sources

PMID: 17822310
PubMed | Europe PMC

Search this title in

Google Scholar