Re: Photobleach (FRAP) problem in Zeiss 510Meta
Posted by
Julio Vazquez on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Photobleach-FRAP-problem-in-Zeiss-510Meta-tp5038048p5038534.html
Guosheng,
I think we are a bit confused as to whether you are unable to run the Photobleaching procedure with the LSM software, or whether you are actually not seeing bleaching. The methods I sent you explain how we run the method. you can see whether the procedure is working because you should see the laser scanning the sample during the bleach procedure (with no image collected at the same time).
To find out the best conditions for bleaching, it is best to bleach a small region, so you can photobleach quickly and start imaging very soon afterwards (especially if you incorporate the bleach routine in your time lapse). Yu can also just use high zoom and high laser power and run a time lapse over a very small region (that is acquire maybe 50 frames without delay in between). You should see intensity drop as you keep imaging (and you will how many iterations are needed for e.g. 50% bleaching). You can then reduce laser power to normal levels and repeat time lapse to see if you have recovery. Once you have determined that, you can set your bleaching/imaging parameters accordingly.
On May 11, 2010, at 2:04 PM, G.Liu wrote:
The sample is tobacco leaf (live).
And I have contacted Zeiss specialist. Due to no internet hookup for the
confocal computer, I did not expect a remote LIVE online trouble shooting.
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