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
Volume6089
DOIs
StatePublished - 2006
Externally publishedYes
EventMultiphoton Microscopy in the Biomedical Sciences VI - San Jose, CA, United States
Duration: Jan 22 2006Jan 24 2006

Other

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

Fingerprint

Fluorescence
Spectroscopy
Color
color
fluorescence
Fluorophores
spectroscopy
bivariate analysis
Detectors
moments
detectors
occupation
Analytical models
Luminance
brightness
Photons
Dyes
Lighting
dyes
illumination

Keywords

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

ASJC Scopus subject areas

  • Electrical and Electronic Engineering
  • Condensed Matter Physics

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 (Vol. 6089). [60890J] https://doi.org/10.1117/12.659642

Cumulant analysis in two-color fluorescence fluctuation spectroscopy. / Wu, Bin; Chen, Y.; Müller, J. D.

Proceedings of SPIE - The International Society for Optical Engineering. Vol. 6089 2006. 60890J.

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

Wu, B, Chen, Y & Müller, JD 2006, Cumulant analysis in two-color fluorescence fluctuation spectroscopy. in Proceedings of SPIE - The International Society for Optical Engineering. vol. 6089, 60890J, Multiphoton Microscopy in the Biomedical Sciences VI, San Jose, CA, United States, 1/22/06. https://doi.org/10.1117/12.659642
Wu B, Chen Y, Müller JD. Cumulant analysis in two-color fluorescence fluctuation spectroscopy. In Proceedings of SPIE - The International Society for Optical Engineering. Vol. 6089. 2006. 60890J https://doi.org/10.1117/12.659642
Wu, Bin ; Chen, Y. ; Müller, J. D. / Cumulant analysis in two-color fluorescence fluctuation spectroscopy. Proceedings of SPIE - The International Society for Optical Engineering. Vol. 6089 2006.
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