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Two-dimensional fluorescence intensity distribution analysis: theory and applications.

Publication ,  Journal Article
Kask, P; Palo, K; Fay, N; Brand, L; Mets, U; Ullmann, D; Jungmann, J; Pschorr, J; Gall, K
Published in: Biophysical journal
April 2000

A method of sample analysis is presented which is based on fitting a joint distribution of photon count numbers. In experiments, fluorescence from a microscopic volume containing a fluctuating number of molecules is monitored by two detectors, using a confocal microscope. The two detectors may have different polarizational or spectral responses. Concentrations of fluorescent species together with two specific brightness values per species are determined. The two-dimensional fluorescence intensity distribution analysis (2D-FIDA), if used with a polarization cube, is a tool that is able to distinguish fluorescent species with different specific polarization ratios. As an example of polarization studies by 2D-FIDA, binding of 5'-(6-carboxytetramethylrhodamine) (TAMRA)-labeled theophylline to an anti-theophylline antibody has been studied. Alternatively, if two-color equipment is used, 2D-FIDA can determine concentrations and specific brightness values of fluorescent species corresponding to individual labels alone and their complex. As an example of two-color 2D-FIDA, binding of TAMRA-labeled somatostatin-14 to the human type-2 high-affinity somatostatin receptors present in stained vesicles has been studied. The presented method is unusually accurate among fluorescence fluctuation methods. It is well suited for monitoring a variety of molecular interactions, including receptors and ligands or antibodies and antigens.

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Published In

Biophysical journal

DOI

EISSN

1542-0086

ISSN

0006-3495

Publication Date

April 2000

Volume

78

Issue

4

Start / End Page

1703 / 1713

Related Subject Headings

  • Theophylline
  • Somatostatin
  • Rhodamines
  • Receptors, Somatostatin
  • Photons
  • Models, Theoretical
  • Microscopy, Fluorescence
  • Microscopy, Confocal
  • Humans
  • Fluorescent Dyes
 

Citation

APA
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Kask, P., Palo, K., Fay, N., Brand, L., Mets, U., Ullmann, D., … Gall, K. (2000). Two-dimensional fluorescence intensity distribution analysis: theory and applications. Biophysical Journal, 78(4), 1703–1713. https://doi.org/10.1016/s0006-3495(00)76722-1
Kask, P., K. Palo, N. Fay, L. Brand, U. Mets, D. Ullmann, J. Jungmann, J. Pschorr, and K. Gall. “Two-dimensional fluorescence intensity distribution analysis: theory and applications.Biophysical Journal 78, no. 4 (April 2000): 1703–13. https://doi.org/10.1016/s0006-3495(00)76722-1.
Kask P, Palo K, Fay N, Brand L, Mets U, Ullmann D, et al. Two-dimensional fluorescence intensity distribution analysis: theory and applications. Biophysical journal. 2000 Apr;78(4):1703–13.
Kask, P., et al. “Two-dimensional fluorescence intensity distribution analysis: theory and applications.Biophysical Journal, vol. 78, no. 4, Apr. 2000, pp. 1703–13. Epmc, doi:10.1016/s0006-3495(00)76722-1.
Kask P, Palo K, Fay N, Brand L, Mets U, Ullmann D, Jungmann J, Pschorr J, Gall K. Two-dimensional fluorescence intensity distribution analysis: theory and applications. Biophysical journal. 2000 Apr;78(4):1703–1713.
Journal cover image

Published In

Biophysical journal

DOI

EISSN

1542-0086

ISSN

0006-3495

Publication Date

April 2000

Volume

78

Issue

4

Start / End Page

1703 / 1713

Related Subject Headings

  • Theophylline
  • Somatostatin
  • Rhodamines
  • Receptors, Somatostatin
  • Photons
  • Models, Theoretical
  • Microscopy, Fluorescence
  • Microscopy, Confocal
  • Humans
  • Fluorescent Dyes