Assessment of membrane fluidity in individual yeast cells by Laurdan generalised polarisation and multi-photon scanning fluorescence microscopy

Learmonth, Robert P. and Gratton, Enrico (2002) Assessment of membrane fluidity in individual yeast cells by Laurdan generalised polarisation and multi-photon scanning fluorescence microscopy. In: Fluorescence spectroscopy, imaging and probes: new tools in chemical, physical and life sciences. Fluorescence: Methods and Applications (2). Springer-Verlag, Heidelberg, Germany, pp. 241-252. ISBN 3-540-42768-6

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Abstract

Here we describe techniques that we developed for monitoring membrane fluidity of individual yeast cells during environmental adaptation and physiological changes. Multi-photon scanning fluorescence microscopy using laurdan as a membrane probe enables determination whether fluidity changes seen by spectroscopy reflect universal responses or changes only of sub-populations. Yeast membranes are a primary site of environmental response and adaptation. Using fluorescence spectroscopy with DPH polarization and laurdan Generalized Polarization (GP), we previously found rapid 'average' membrane fluidity modulation in yeast populations during growth and in response to nutrients or environmental stresses. To determine whether such responses reflect all cells we conducted the first multi-photon scanning fluorescence microscopy study of yeasts, measuring Laurdan GP. We assessed membrane fluidity responses of individual yeasts related to growth phase, heat stress and ethanol stress. Average fluidity decreased as cultures aged, however the decreased fluidity was due in some cases to an increasing proportion of uniformly low fluidity (high GP) cells, which were shown by vital dye to be dead. When yeasts were heat stressed, the mean Laurdan GP increased in all cells, thus the entire population evidenced damage (viz. decreased membrane fluidity) to the same degree. On the other hand, with ethanol stress fluidity increased (GP decreased) on exposure of cells. All cells were affected although not to the same degree, and with variable recovery. The recovery assessed from GP microscopy was highly variable, and greater by that seen by spectroscopy.


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Item Type: Book Chapter (Commonwealth Reporting Category B)
Refereed: Yes
Item Status: Live Archive
Additional Information: Author's version deposited with blanket permission of publisher. Print copy not held USQ Library (Order cancelled 14/3/2008 as out of print). A collection of papers documenting the 7th Conference on 'Methods and Applications of Fluorescence (MAF): Spectroscopy, Imaging and Probes'. This conference was held in September 2001.
Depositing User: Assoc Prof Robert Learmonth
Faculty / Department / School: Historic - Faculty of Sciences - Department of Biological and Physical Sciences
Date Deposited: 08 Nov 2007 07:07
Last Modified: 10 Oct 2013 02:30
Uncontrolled Keywords: yeast; membrane; fluidity; 2-photon; microscopy; laser; scanning; Laurdan; generalized polarization
Fields of Research (FOR2008): 06 Biological Sciences > 0601 Biochemistry and Cell Biology > 060110 Receptors and Membrane Biology
02 Physical Sciences > 0205 Optical Physics > 020599 Optical Physics not elsewhere classified
06 Biological Sciences > 0605 Microbiology > 060599 Microbiology not elsewhere classified
Socio-Economic Objective (SEO2008): E Expanding Knowledge > 97 Expanding Knowledge > 970106 Expanding Knowledge in the Biological Sciences
URI: http://eprints.usq.edu.au/id/eprint/3326

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