Posted by
Dan Stevens-2 on
URL: http://confocal-microscopy-list.275.s1.nabble.com/XYZ-drifts-giving-advice-and-looking-for-advice-tp5977803p6002724.html
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Hi Daniel,
There has been some good feedback here, but I wanted to add two bits that I
did not see posted.
1 - I have seen a good number of people equilibrate their microscope and then
drop the sample in and begin imaging. The sample needs to equilibrate. You
can do a nice job of isolating whether the drift is associated with sample (most
often in my experience) or stand by using a fixed sample loosely fitted in the
carrier. I have used PSF beads, so that the original plane of focus is easily
defined and the degree of drift can be found at the end of the time course.
2 - In performing heat shock experiments, I have noted xy drift. These
experiments were done with perfusion, with the stage incubator kept at a
constant, over time periods where I expect there was effectively no heat
transfer beyond that carried by the bulk flow. The use of a hardware based
autofocus device (zeiss definite focus), allowed us to measure the degree and
rate of z-focus drift, but a bit to my surprise we also saw significant xy drift.
To my best recollection the heat shock was 5 degrees rise over 2 minutes, the
resulting z drift (measured by degree of system compensation to maintain the
plane) was 4um, and the xy drift was ~10um. The xy drift reversed when we
dropped the temp back.
Thought that might be of some interest to you, given you saw drift in xy as
well as z.
regards,
Dan Stevens, PhD
Application Specialist
Carl Zeiss Canada
*Strong commercial interest