Please use this identifier to cite or link to this item: https://doi.org/10.1529/biophysj.105.074120
Title: Simultaneous multicolor fluorescence cross-correlation spectroscopy to detect higher order molecular interactions using single wavelength laser excitation
Authors: Ling, C.H.
Gösch, M.
Lasser, T.
Wohland, T. 
Issue Date: 2006
Citation: Ling, C.H., Gösch, M., Lasser, T., Wohland, T. (2006). Simultaneous multicolor fluorescence cross-correlation spectroscopy to detect higher order molecular interactions using single wavelength laser excitation. Biophysical Journal 91 (2) : 715-727. ScholarBank@NUS Repository. https://doi.org/10.1529/biophysj.105.074120
Abstract: Fluorescence cross-correlation spectroscopy is a powerful method for the study of molecular interactions and dynamics in solution and even in living cells. Usually, in the optical setup, either two laser beams have to be superimposed in their respective confocal volumes or two-photon excitation is used for a dual-color detection system. It has been shown recently that fluorescence cross correlation can be achieved with spectrally similar fluorophores using single wavelength excitation fluorescence cross-correlation spectroscopy (SW-FCCS). In this study, we show that SW-FCCS allows the simultaneous excitation of up to three fluorophores in which the cross correlation of their fluctuation signals is detected separately in three detection channels. The experimental and theoretical model to describe triple pairwise cross correlations incorporating cross talk and possible changes in emission characteristics such as quenching upon binding are outlined. The effectiveness of SW-FCCS to detect binding of three interacting partners is experimentally verified with a standard ligand-receptor model, biotin-streptavidin, where differently labeled biotin ligands and their binding to a third-color labeled streptavidin are studied. The cross-correlation amplitudes and their changes with stoichiometric binding are analyzed and the upper limits of dissociation constants are determined. Performed with appropriate negative controls, SW-FCCS can determine interaction patterns between ligands and receptors. © 2006 by the Biophysical Society.
Source Title: Biophysical Journal
URI: http://scholarbank.nus.edu.sg/handle/10635/76978
ISSN: 00063495
DOI: 10.1529/biophysj.105.074120
Appears in Collections:Staff Publications

Show full item record
Files in This Item:
There are no files associated with this item.

SCOPUSTM   
Citations

20
checked on Sep 10, 2018

WEB OF SCIENCETM
Citations

30
checked on Sep 3, 2018

Page view(s)

18
checked on Jun 22, 2018

Google ScholarTM

Check

Altmetric


Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.