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Supercritical angle fluorescence correlation spectroscopy


Ries, J; Ruckstuhl, T; Verdes, D; Schwille, P (2008). Supercritical angle fluorescence correlation spectroscopy. Biophysical Journal, 94(1):221-229.

Abstract

We explore the potential of a supercritical angle (SA) objective for fluorescence correlation spectroscopy (FCS). This novel microscope objective combines tight focusing by an aspheric lens with strong axial confinement of supercritical angle fluorescence collection by a parabolic mirror lens, resulting in a small detection volume. The tiny axial extent of the detection volume features an excellent surface sensitivity, as is demonstrated by diffusion measurements in model membranes with an excess of free dye in solution. All SA-FCS measurements are directly compared to standard confocal FCS, demonstrating a clear advantage of SA-FCS, especially for diffusion measurements in membranes. We present an extensive theoretical framework that allows for accurate and quantitative evaluation of the SA-FCS correlation curves.

Abstract

We explore the potential of a supercritical angle (SA) objective for fluorescence correlation spectroscopy (FCS). This novel microscope objective combines tight focusing by an aspheric lens with strong axial confinement of supercritical angle fluorescence collection by a parabolic mirror lens, resulting in a small detection volume. The tiny axial extent of the detection volume features an excellent surface sensitivity, as is demonstrated by diffusion measurements in model membranes with an excess of free dye in solution. All SA-FCS measurements are directly compared to standard confocal FCS, demonstrating a clear advantage of SA-FCS, especially for diffusion measurements in membranes. We present an extensive theoretical framework that allows for accurate and quantitative evaluation of the SA-FCS correlation curves.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:07 Faculty of Science > Department of Chemistry
Dewey Decimal Classification:540 Chemistry
Language:English
Date:2008
Deposited On:24 Jan 2009 16:48
Last Modified:06 Dec 2017 17:20
Publisher:Elsevier
ISSN:0006-3495
Free access at:PubMed ID. An embargo period may apply.
Publisher DOI:https://doi.org/10.1529/biophysj.107.115998
PubMed ID:17827221

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