Please use this identifier to cite or link to this item: https://hdl.handle.net/1959.11/10034
Title: Single point FCS and number and brightness analysis on a commercial confocal laser scanning microscope with analog detectors (Nikon C1)
Contributor(s): Salvemini, Iyrri (author); Gratton, Enrico (author); Moens, Pierre  (author)orcid 
Publication Date: 2009
Handle Link: https://hdl.handle.net/1959.11/10034
Abstract: Fluorescence Correlation Spectroscopy is a technique developed in the early 1970s to measure chemical reaction rates, diffusion coefficient and photo physical processes. Commonly single point FCS data are obtained on sophisticated FCS instruments operating in the photon counting mode. Here we show that accurate FCS data can be obtained on a low cost commercial confocal laser scanning microscope without any modifications. Using the Nikon C1, we measured the diffusion coefficient and the concentration of Rhodamine B in solution for concentrations ranging from 5 nM to 280 nM. We also determined the diffusion coefficient of two different labeled lipid analogs (1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate and BODIPY TMR phosphatidylinositol (4,5) bisphosphate) incorporated into the membrane of giant unilamellar vesicles. The results obtained for these lipid analogs are in good agreementwith previously published data. Although single point FCS is possible with analog detectors the photon counting histogramanalysis is not. Number and brightness analysis is an alternative to PCH to determine the aggregation state of molecules. We show that the molecules incorporated into the GUV membrane are uniformly distributed into the membrane of the GUV. Finally, we highlight the fact that the actual proportion of labeled lipid analogs incorporated into the membrane of the giant unilamellar vesicle (formed by the electroformation method) is significantly different than the proportion of these lipids in the organic solvent stock solution.
Publication Type: Conference Publication
Conference Details: ASB 2009: 33rd Annual Meeting of the Australian Society for Biophysics, Ballarat, Australia, 29th November - 2nd December, 2009
Grant Details: NHMRC/568301
Source of Publication: 33rd Annual Meeting of the Australian Society for Biophysics Programme, p. 20-20
Publisher: Australian Society for Biophysics
Place of Publication: Australia
Fields of Research (FoR) 2008: 029901 Biological Physics
Socio-Economic Objective (SEO) 2008: 970106 Expanding Knowledge in the Biological Sciences
970110 Expanding Knowledge in Technology
HERDC Category Description: E3 Extract of Scholarly Conference Publication
Publisher/associated links: http://biophysics.chemistry.unimelb.edu.au/ASB_Ballarat/ASB_2009_files/ASB2009_program.pdf
Appears in Collections:Conference Publication

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