Fluorescence Correlation Spectroscopy at Micromolar Concentrations without Optical Nanoconfinement
Abstract
Fluorescence correlation spectroscopy (FCS) is an important technique for studying biochemical interactions dynamically that may be used in vitro and in cell-based studies. It is generally claimed that FCS may only be used at nM concentrations. We show that this general consensus is incorrect and that the limitation to nM concentrations is not fundamental but due to detector limits as well as laser fluctuations. With a high count rate detector system and applying laser fluctuation corrections, we demonstrate FCS measurements up to 38 μM with the same signal-to-noise as at lower concentrations. Optical nanoconfinement approaches previously used to increase the concentration range of FCS are not necessary, and further increases above 38 μM may be expected using detectors and detector arrays with higher saturation rates and better laser fluctuation corrections. This approach greatly widens the possibilities of dynamic measurements of biochemical interactions using FCS at physiological concentrations.
- Authors:
-
- Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
- Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Univ. of California, Davis, CA (United States). Dept. of Radiation Oncology
- Publication Date:
- Research Org.:
- Lawrence Livermore National Laboratory (LLNL), Livermore, CA (United States)
- Sponsoring Org.:
- USDOE
- OSTI Identifier:
- 1169272
- Report Number(s):
- LLNL-JRNL-653521
Journal ID: ISSN 1520-6106
- Grant/Contract Number:
- AC52-07NA27344; 11-ERD-037
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Journal of Physical Chemistry. B, Condensed Matter, Materials, Surfaces, Interfaces and Biophysical Chemistry
- Additional Journal Information:
- Journal Volume: 118; Journal Issue: 32; Journal ID: ISSN 1520-6106
- Publisher:
- American Chemical Society
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 37 INORGANIC, ORGANIC, PHYSICAL AND ANALYTICAL CHEMISTRY; 59 BASIC BIOLOGICAL SCIENCES; Fluorescence correlation spectroscopy (FCS)
Citation Formats
Laurence, Ted A., Ly, Sonny, Bourguet, Feliza, Fischer, Nicholas O., and Coleman, Matthew A. Fluorescence Correlation Spectroscopy at Micromolar Concentrations without Optical Nanoconfinement. United States: N. p., 2014.
Web. doi:10.1021/jp505881z.
Laurence, Ted A., Ly, Sonny, Bourguet, Feliza, Fischer, Nicholas O., & Coleman, Matthew A. Fluorescence Correlation Spectroscopy at Micromolar Concentrations without Optical Nanoconfinement. United States. https://doi.org/10.1021/jp505881z
Laurence, Ted A., Ly, Sonny, Bourguet, Feliza, Fischer, Nicholas O., and Coleman, Matthew A. Thu .
"Fluorescence Correlation Spectroscopy at Micromolar Concentrations without Optical Nanoconfinement". United States. https://doi.org/10.1021/jp505881z. https://www.osti.gov/servlets/purl/1169272.
@article{osti_1169272,
title = {Fluorescence Correlation Spectroscopy at Micromolar Concentrations without Optical Nanoconfinement},
author = {Laurence, Ted A. and Ly, Sonny and Bourguet, Feliza and Fischer, Nicholas O. and Coleman, Matthew A.},
abstractNote = {Fluorescence correlation spectroscopy (FCS) is an important technique for studying biochemical interactions dynamically that may be used in vitro and in cell-based studies. It is generally claimed that FCS may only be used at nM concentrations. We show that this general consensus is incorrect and that the limitation to nM concentrations is not fundamental but due to detector limits as well as laser fluctuations. With a high count rate detector system and applying laser fluctuation corrections, we demonstrate FCS measurements up to 38 μM with the same signal-to-noise as at lower concentrations. Optical nanoconfinement approaches previously used to increase the concentration range of FCS are not necessary, and further increases above 38 μM may be expected using detectors and detector arrays with higher saturation rates and better laser fluctuation corrections. This approach greatly widens the possibilities of dynamic measurements of biochemical interactions using FCS at physiological concentrations.},
doi = {10.1021/jp505881z},
journal = {Journal of Physical Chemistry. B, Condensed Matter, Materials, Surfaces, Interfaces and Biophysical Chemistry},
number = 32,
volume = 118,
place = {United States},
year = {Thu Aug 14 00:00:00 EDT 2014},
month = {Thu Aug 14 00:00:00 EDT 2014}
}
Web of Science
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Works referencing / citing this record:
State-of-the-Art Fluorescence Fluctuation-Based Spectroscopic Techniques for the Study of Protein Aggregation
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High photon count rates improve the quality of super-resolution fluorescence fluctuation spectroscopy
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