help button home button Biophys. J.
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Originally published as Biophys J. BioFAST on May 13, 2005.
doi:10.1529/biophysj.105.060749
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
biophysj.105.060749v1
89/2/1288    most recent
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Skinner, J. P.
Right arrow Articles by Müller, J. D.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Skinner, J. P.
Right arrow Articles by Müller, J. D.
Biophysical Journal 89:1288-1301 (2005)
© 2005 The Biophysical Society

Position-Sensitive Scanning Fluorescence Correlation Spectroscopy

Joseph P. Skinner, Yan Chen and Joachim D. Müller

School of Physics and Astronomy, University of Minnesota, Minneapolis, Minnesota 55455

Correspondence: Address reprint requests to Joseph P. Skinner, University of Minnesota, School of Physics and Astronomy, 116 Church St., SE Minneapolis, MN 55455. Tel.: 612-624-6045; Fax: 612-624-4578; E-mail: josephs{at}physics.umn.edu.

Fluorescence correlation spectroscopy (FCS) uses a stationary laser beam to illuminate a small sample volume and analyze the temporal behavior of the fluorescence fluctuations within the stationary observation volume. In contrast, scanning FCS (SFCS) collects the fluorescence signal from a moving observation volume by scanning the laser beam. The fluctuations now contain both temporal and spatial information about the sample. To access the spatial information we synchronize scanning and data acquisition. Synchronization allows us to evaluate correlations for every position along the scanned trajectory. We use a circular scan trajectory in this study. Because the scan radius is constant, the phase angle is sufficient to characterize the position of the beam. We introduce position-sensitive SFCS (PSFCS), where correlations are calculated as a function of lag time and phase. We present the theory of PSFCS and derive expressions for diffusion, diffusion in the presence of flow, and for immobilization. To test PSFCS we compare experimental data with theory. We determine the direction and speed of a flowing dye solution and the position of an immobilized particle. To demonstrate the feasibility of the technique for applications in living cells we present data of enhanced green fluorescent protein measured in the nucleus of COS cells.




This article has been cited by other articles:


Home page
 Biophys. JHome page
B. Wu, Y. Chen, and J. D. Muller
Fluorescence Correlation Spectroscopy of Finite-Sized Particles
Biophys. J., April 1, 2008; 94(7): 2800 - 2808.
[Abstract] [Full Text] [PDF]

Home page
 Biophys. JHome page
Z. Petrasek and P. Schwille
Precise Measurement of Diffusion Coefficients using Scanning Fluorescence Correlation Spectroscopy
Biophys. J., February 15, 2008; 94(4): 1437 - 1448.
[Abstract] [Full Text] [PDF]

Home page
 Biophys. JHome page
D. R. Sisan, R. Arevalo, C. Graves, R. McAllister, and J. S. Urbach
Spatially Resolved Fluorescence Correlation Spectroscopy Using a Spinning Disk Confocal Microscope
Biophys. J., December 1, 2006; 91(11): 4241 - 4252.
[Abstract] [Full Text] [PDF]

Home page
 Biophys. JHome page
J. Ries and P. Schwille
Studying Slow Membrane Dynamics with Continuous Wave Scanning Fluorescence Correlation Spectroscopy
Biophys. J., September 1, 2006; 91(5): 1915 - 1924.
[Abstract] [Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Copyright © 2005 by the Biophysical Society.