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21 June 2004 Dynamic and static fluorescence anisotropy in biological microscopy (rFLIM and emFRET)
Thomas M. Jovin, Diane S. Lidke, Janine N. Post
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Proceedings Volume 5323, Multiphoton Microscopy in the Biomedical Sciences IV; (2004) https://doi.org/10.1117/12.528469
Event: Biomedical Optics 2004, 2004, San Jose, CA, United States
Abstract
Fluorescence anisotropy, a measure of the polarization state of fluorescence emission, is a sensitive measure of molecular rotational motion and of resonance energy transfer (RET). We report here the formalism and application of dynamic and static fluorescence anisotropy measurements primarily intended for implementation in imaging systems. These include confocal lasre scanning microscopes (CLSM) as well as wide-field instruments, in the latter case adapted for anisotropy-based dynamic frequency domain fluorescence lifetime imaging microscopy (FLIM), a method we denote as rFLIM. Anisotropy RET is one of the modalities used for fluorescence RET (FRET) determinations of the association, and proximity of cellular proteins in vivo. A requirement is the existence of intrinsic or extrinsic probes exhibiting homotransfer FRET (in our nomenclature, energy migration or emFRET) between like fluorophores. This phenomenon is particularly useful in studies of the activation and processing of transmembrane receptor tyrosine kinases involved in signal transduction and expressed as fusions with Visible Fluorescence Proteins (VFPs).
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Thomas M. Jovin, Diane S. Lidke, and Janine N. Post "Dynamic and static fluorescence anisotropy in biological microscopy (rFLIM and emFRET)", Proc. SPIE 5323, Multiphoton Microscopy in the Biomedical Sciences IV, (21 June 2004); https://doi.org/10.1117/12.528469
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KEYWORDS
Anisotropy
Venus
Luminescence
Proteins
Fluorescence resonance energy transfer
Fluorescence anisotropy
Microscopy
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