Open Access3D fluorescence anisotropy imaging using selective plane illumination microscopy
Author(s) -
Per Niklas Hedde,
Suman Ranjit,
Enrico Gratton
Publication year - 2015
Publication title -
optics express
Language(s) - Uncategorized
Resource type - Journals
SCImago Journal Rank - 1.394
H-Index - 271
ISSN - 1094-4087
DOI - 10.1364/oe.23.022308
Subject(s) - microscopy , optics , light sheet fluorescence microscopy , photobleaching , anisotropy , materials science , fluorescence lifetime imaging microscopy , fluorescence microscope , photoactivated localization microscopy , microscope , fluorescence , confocal microscopy , confocal , optical sectioning , biological imaging , super resolution microscopy , scanning confocal electron microscopy , physics
Fluorescence anisotropy imaging is a popular method to visualize changes in organization and conformation of biomolecules within cells and tissues. In such an experiment, depolarization effects resulting from differences in orientation, proximity and rotational mobility of fluorescently labeled molecules are probed with high spatial resolution. Fluorescence anisotropy is typically imaged using laser scanning and epifluorescence-based approaches. Unfortunately, those techniques are limited in either axial resolution, image acquisition speed, or by photobleaching. In the last decade, however, selective plane illumination microscopy has emerged as the preferred choice for three-dimensional time lapse imaging combining axial sectioning capability with fast, camera-based image acquisition, and minimal light exposure. We demonstrate how selective plane illumination microscopy can be utilized for three-dimensional fluorescence anisotropy imaging of live cells. We further examined the formation of focal adhesions by three-dimensional time lapse anisotropy imaging of CHO-K1 cells expressing an EGFP-paxillin fusion protein.