membrane potential lab

[Jon Monroe]

Plant-eders,

I have the notes for a lab that I would like to try to adapt for a 
Plant Physiology course in which students isolate membrane vesicles 
enriched in tonoplast or plasma membrane and use fluorescent probes 
to detect the formation of a pH gradient or an electrical potential. 
The lab was written by Heven Sze at the U of Maryland and is (I 
believe) based on her paper: Randall SK, Sze H (1986) Properties of 
the partially purified tonoplast H+-pumping ATPase from oat roots. J. 
Biol. Chem. 261: 1364-1371.

A key aspect of the lab is the use of acridine orange and oxonol V 
which fluoresce.  Unfortunately we don't have lots of fluorescence 
spectrophotometers...  Are there any alternatives to these compounds 
which could be used with UV/Vis spectrophotometers?  I wonder if the 
signal to noise ratios using visible compounds are just too low for 
these sorts of measurements.  Do any of you have a similar lab 
exercise that you could share?  Thanks!

Jon

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  Jonathan D. Monroe           Associate Professor
  Department of Biology       office: 540-568-6649
  MSC 7801                       lab: 540-568-6045
  James Madison University       fax: 540-568-3333
  Harrisonburg, VA 22807   email: monroejd at jmu.edu
  http://csm.jmu.edu/biology/monroejd/jmonroe.html
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