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Ignatius, Mike on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Strange-artifact-in-z-series-tp592967p592972.html
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http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocalIt is not uncommon to de-quench mitochondrial dyes with light. As the dye bleaches, it is no longer is available to quenches others around it. Eventually an optimum is reached, where the mitochondria are now brighter - or perfectly labeled. Same thing can happen with dyes on antibodies.
One way to test is to reverse the stack order or to label with less dye. But sounds like a feature, not a flaw here?
Mike Ignatius
Molecular Probes/Invitrogen
-----Original Message-----
From: Confocal Microscopy List [mailto:
[hidden email]] On Behalf Of Pedro Camello
Sent: Tuesday, May 06, 2008 3:44 PM
To:
[hidden email]
Subject: Strange artifact in z series
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http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocalHi all,
while performing z sections of mitochondria-stained living cells, I´m
finding that the bottom part of the cell displays a brighter mitochondria
fluoresecence, showing more contrast respect to cytosol than mitochondria in
the top part of the cell. My system scans z from bottom to top. If it were
simple bleaching due to laser light passing through the cell, I should
observe that the bottom mitochondria are also less stained in a second
series, but that´s not the case (the difference between top and bottom still
there)
Any idea to explain it?
Thanks
Pedro