Advances in polarized fluorescence depletion measurement of cell membrane protein rotation

B. George Barisas; N. A. Rahman; Thomas Londo; J. R. Herman; Deborah A. Roess

doi:10.1117/12.44208

1 June 1991 Advances in polarized fluorescence depletion measurement of cell membrane protein rotation

B. George Barisas, N. A. Rahman, Thomas Londo, J. R. Herman, Deborah A. Roess

Proceedings Volume 1432, Biomolecular Spectroscopy II; (1991) https://doi.org/10.1117/12.44208
Event: Optics, Electro-Optics, and Laser Applications in Science and Engineering, 1991, Los Angeles, CA, United States

Abstract

The laser microscopic method of polarized fluorescence depletion (PFD) has permitted the rotational dynamics of functional membrane proteins to be measured on single cells under physiological conditions. This provides information comparable to that obtained by the older technique of time-resolved phosphorescence anisotropy (TPA) but with a greater than 1000- fold reduction in sample requirements. A new cuvette implementation of PFD methods, together with a data analysis model appropriate to this new experimental geometry, permits small volumes of dilute cell suspensions to be examined in their entirety. Data can thus be obtained at rates comparable to TPA. The instrument can also be used for TPA measurements and this permits direct comparison of PFD and TPA results obtained under otherwise identical conditions. In addition, a new photomultiplier gating device and system timing strategy reduce the minimum observable rotational correlation times to < 2 microsecond(s) ec, a 10-fold improvement over previous systems. These speed improvements have been examined in studies of BSA rotation in glycerol solution.

Citation Download Citation

B. George Barisas, N. A. Rahman, Thomas Londo, J. R. Herman, and Deborah A. Roess "Advances in polarized fluorescence depletion measurement of cell membrane protein rotation", Proc. SPIE 1432, Biomolecular Spectroscopy II, (1 June 1991); https://doi.org/10.1117/12.44208

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