Cumulant analysis in two-color fluorescence fluctuation spectroscopy

Bin Wu, Y. Chen, J. D. Müller

Research output: Chapter in Book/Report/Conference proceedingConference contribution

Abstract

Fluorescence Fluctuation Spectroscopy (FFS) studies the fluctuating fluorescent signal from a small illumination volume and extracts the concentration and dynamical information of fluorophores. Detecting the fluorescence in two detector channels introduces the possibility of differentiating the fluorophores based on color. We introduce bivariate cumulant analysis for Two-Color Fluorescence Fluctuation Spectroscopy and derive an analytical expression for the bivariate factorial cumulants of photon counts at arbitrary sampling times. Fits of the data to the analytical model determine the brightness of each channel, occupation number and diffusion time of each fluorescent species. The statistical accuracy of each cumulant is described by its variance, which we calculate by the moments-of-moments technique. The theory is experimentally verified using model dye system. We also performed first experiments in living cells, and develop a model that takes nonideal detector effects into account. This technique is useful for optimizing the spectroscopic separation of heterogeneous biological samples by FFS.

Original languageEnglish (US)
Title of host publicationProceedings of SPIE - The International Society for Optical Engineering
DOIs
StatePublished - May 8 2006
Externally publishedYes
EventMultiphoton Microscopy in the Biomedical Sciences VI - San Jose, CA, United States
Duration: Jan 22 2006Jan 24 2006

Publication series

NameProceedings of SPIE - The International Society for Optical Engineering
Volume6089
ISSN (Print)0277-786X

Other

OtherMultiphoton Microscopy in the Biomedical Sciences VI
CountryUnited States
CitySan Jose, CA
Period1/22/061/24/06

Keywords

  • Cumulant
  • Fluorescence Fluctuation Spectroscopy
  • Fluorescence correlation spectroscopy
  • Photon statistics
  • Two-Color
  • Two-photon microscopy

ASJC Scopus subject areas

  • Electronic, Optical and Magnetic Materials
  • Condensed Matter Physics
  • Computer Science Applications
  • Applied Mathematics
  • Electrical and Electronic Engineering

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  • Cite this

    Wu, B., Chen, Y., & Müller, J. D. (2006). Cumulant analysis in two-color fluorescence fluctuation spectroscopy. In Proceedings of SPIE - The International Society for Optical Engineering [60890J] (Proceedings of SPIE - The International Society for Optical Engineering; Vol. 6089). https://doi.org/10.1117/12.659642