Re: Use of a fluorescent organelle in determining a PSF

Posted by Sarah Kefayati on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Coverslip-tp592448p592458.html

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Dear Fred:

 

You may find this article useful,they also extract the PSF from the spherical shape objects from the sample.

 

http://www.biophysj.org/cgi/reprint/85/6/3991

 

 

Sarah Kefayati

Biophysics MSc student

Brock university

St. Catharines

CANADA

 

On Fri, Feb 22, 2008 at 4:54 PM, Fred Mast <[hidden email]> wrote:

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Hello all,

I would like some input into an idea I have for obtaining a PSF. I
study peroxisomes using yeast as a model system for understanding how
they are created. In yeast, peroxisomes have a well characterized
morphology, being spherical organelles with a diameter of 100 to
200nm. Other than this size variability I think they are excellent
candidates for obtaining a PSF as they can be easily, fluorescently
labelled (by targeting fluorescent protein chimeras to their matrix),
are similar in size to what is typically used to obtain PSF's, and are
"embedded" in the sample. I do a lot of live cell imaging, using a
LSM510 Meta and am always looking for ways to improve my system. As a
result most of my images are fairly noisy and I rely on deconvolution
to remove the noise, and improve contrast and resolution. My initial
attempts at using peroxisomes for this purpose have provided me with a
PSF that is slightly different from what I obtain with fluorescent
beads (the peroxisome derived PSF is less symmetrical) and provides,
in my estimation, a more realistic result. Your thoughts and concerns
on this idea would be most welcome.

Fred

Fred D. Mast
Department of Cell Biology
Medical Sciences Building Room 5-14
University of Alberta
Edmonton, Alberta, T6G 2H7
Canada

Tel: [phone]
[hidden email]