1 April 2002 Measurement of cell surface protein dynamics by two-photon image correlation spectroscopy and image cross-correlation spectroscopy
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Proceedings Volume 4633, Commercial and Biomedical Applications of Ultrafast and Free-Electron Lasers; (2002); doi: 10.1117/12.461365
Event: High-Power Lasers and Applications, 2002, San Jose, California, United States
Abstract
Advances in laser-scanning microscopy and the advent of confocal microscopy permitted the development of image correlation spectroscopy (ICS). ICS is an imaging analog of fluorescence correlation spectroscopy (FCS) optimized for measuring the aggregation state of fluorescently labeled macromolecules on the surface of biological cells. The ICS method entails spatial autocorrelation analysis of fluorescence fluctuations within an image sampled from an area of the sample as well as temporal autocorrelation analysis of fluorescence fluctuations through a time series of images. Together, the spatial/temporal autocorrelation analysis enables measurement of fluorophore concentration, aggregation state and transport properties. ICS was first implemented on a confocal laser-scanning microscope (CLSM) using single photon excitation. More recently we have extended the method for two-photon ICS as well as image cross-correlation spectroscopy (ICCS). ICCS allows measurement of co-localization of non-identical molecules labeled with fluorophores of different emission wavelengths. We present a variety of applications of the ICS and ICCS methods in cellular systems. We will discuss the measurement of the transport and clustering properties of membrane receptors by single photon ICS and two-photon ICCS. As well, we will describe how spatial ICS may be used to quantify the distribution of fluorescently labeled dendritic spines in neurons.
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Paul W. Wiseman, Jeffrey A. Squier, "Measurement of cell surface protein dynamics by two-photon image correlation spectroscopy and image cross-correlation spectroscopy", Proc. SPIE 4633, Commercial and Biomedical Applications of Ultrafast and Free-Electron Lasers, (1 April 2002); doi: 10.1117/12.461365; http://dx.doi.org/10.1117/12.461365
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KEYWORDS
Diffusion

Quantum dots

Spine

Spectroscopy

Imaging spectroscopy

Receptors

Fluorescence correlation spectroscopy

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