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Title: Fluorescence Correlation Spectroscopy at Micromolar Concentrations without Optical Nanoconfinement

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:
 [1] ;  [1] ;  [1] ;  [1] ;  [2]
  1. Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
  2. Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Univ. of California, Davis, CA (United States). Dept. of Radiation Oncology
Publication Date:
OSTI Identifier:
1169272
Report Number(s):
LLNL-JRNL--653521
Journal ID: ISSN 1520-6106
Grant/Contract Number:
AC52-07NA27344; 11-ERD-037
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
Research Org:
Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
Sponsoring Org:
USDOE
Country of Publication:
United States
Language:
English
Subject:
37 INORGANIC, ORGANIC, PHYSICAL AND ANALYTICAL CHEMISTRY; 59 BASIC BIOLOGICAL SCIENCES Fluorescence correlation spectroscopy (FCS)