Document Detail

Separation of translational and rotational contributions in solution studies using fluorescence photobleaching recovery.
MedLine Citation:
PMID:  6518256     Owner:  NLM     Status:  MEDLINE    
Although fluorescence photobleaching recovery (FPR) experiments are usually interpreted in terms of the translational motions of a fluorescently labeled species, rotational motions can also modulate recovery through the cosine-squared laws for dipolar absorption and emission processes. In a complex interacting system, translational and rotational contributions may both be simultaneously present. We show how these contributions can be separated in solution studies using an FPR setup in which (a) the linear polarization of the low-intensity observation beam and the high-intensity photobleaching pulse can be varied independently, and (b) all emitted fluorescent photons are counted equally. The fluorescence recovery signal obtained with the observation beam polarized at the magic angle, 54.7 degrees, from the bleach polarization direction is independent of label orientation, whereas the anisotropy function formed from a combination of parallel and perpendicular polarizations isolates the orientational recovery. The anisotropy function is identical to that in fluorescence correlation spectroscopy and, for rigid-body rotational diffusion, can be expressed as a sum of five exponential terms.
W A Wegener; R Rigler
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Biophysical journal     Volume:  46     ISSN:  0006-3495     ISO Abbreviation:  Biophys. J.     Publication Date:  1984 Dec 
Date Detail:
Created Date:  1985-02-28     Completed Date:  1985-02-28     Revised Date:  2009-11-18    
Medline Journal Info:
Nlm Unique ID:  0370626     Medline TA:  Biophys J     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  787-93     Citation Subset:  IM    
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MeSH Terms
Models, Biological*
Spectrometry, Fluorescence
Grant Support
Reg. No./Substance:

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