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Optics Express

Optics Express

  • Editor: C. Martijn de Sterke
  • Vol. 18, Iss. 20 — Sep. 27, 2010
  • pp: 21225–21237

Measuring and imaging diffusion with multiple scan speed image correlation spectroscopy

Nadine Gröner, Jérémie Capoulade, Christoph Cremer, and Malte Wachsmuth  »View Author Affiliations

Optics Express, Vol. 18, Issue 20, pp. 21225-21237 (2010)

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The intracellular mobility of biomolecules is determined by transport and diffusion as well as molecular interactions and is crucial for many processes in living cells. Methods of fluorescence microscopy like confocal laser scanning microscopy (CLSM) can be used to characterize the intracellular distribution of fluorescently labeled biomolecules. Fluorescence correlation spectroscopy (FCS) is used to describe diffusion, transport and photo-physical processes quantitatively. As an alternative to FCS, spatially resolved measurements of mobilities can be implemented using a CLSM by utilizing the spatio-temporal information inscribed into the image by the scan process, referred to as raster image correlation spectroscopy (RICS). Here we present and discuss an extended approach, multiple scan speed image correlation spectroscopy (msICS), which benefits from the advantages of RICS, i.e. the use of widely available instrumentation and the extraction of spatially resolved mobility information, without the need of a priori knowledge of diffusion properties. In addition, msICS covers a broad dynamic range, generates correlation data comparable to FCS measurements, and allows to derive two-dimensional maps of diffusion coefficients. We show the applicability of msICS to fluorophores in solution and to free EGFP in living cells.

© 2010 OSA

OCIS Codes
(100.2960) Image processing : Image analysis
(180.5810) Microscopy : Scanning microscopy
(300.6500) Spectroscopy : Spectroscopy, time-resolved

ToC Category:

Original Manuscript: July 13, 2010
Manuscript Accepted: August 11, 2010
Published: September 22, 2010

Virtual Issues
Vol. 5, Iss. 14 Virtual Journal for Biomedical Optics

Nadine Gröner, Jérémie Capoulade, Christoph Cremer, and Malte Wachsmuth, "Measuring and imaging diffusion with multiple scan speed image correlation spectroscopy," Opt. Express 18, 21225-21237 (2010)

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