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Hi Kyle,
if your transmitted image is free of astigmatism, then the dichroic must be
responsible for the astigmatism in the reflected path (assuming there are no
other components between the dichoic and the cameras). Try a flat mirror (I
mean really flat, 6mm substrate, lambda/10 specs) instead of the dichroic
and see if the astigmatism is gone.
The new 3mm dichroics have similar specs (lambda/4 to lambda/10), so if it
is mounted properly (it's not about exact angles, but you must not introduce
any stress that may lead to bending), it should work OK. Try another
dichroic, maybe one factory-mounted in a cube.
Also, the closer the dichroic to the camera, the smaller the effects of
curvature. I don't think infinty space is less 'sensitive' to astigmatism.
Quite the opposite - least sensitive should be a position conjugate to image
plane (but then, every piece of dust will be in focus on the camera...).
Best, zdenek
Zdenek Svindrych, Ph.D.
W.M. Keck Center for Cellular Imaging (PLSB 003)
Department of Biology,University of Virginia
409 McCormick Rd, Charlottesville, VA-22904
http://www.kcci.virginia.edu/
tel: 434-982-4869
Annual FRET Workshop (March 6-10, 2017): http://kcci.virginia.edu/workshop-
2017
---------- Původní zpráva ----------
Od: Kyle Douglass <[log in to unmask]>
Komu: [log in to unmask]
Datum: 13. 2. 2017 9:29:31
Předmět: Reflected image -> astigmatism
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Hi everyone,
A couple years ago there was a post on this forum about significant
astigmatism appearing in the reflected path of a custom-built, two
camera TIRF setup.
(http://confocal-microscopy-list.588098.n2.nabble.com/Reflected-image-gt-
astigmatism-td7584402.html)
The original poster had placed a dichroic before a pair of matched tube
lenses to split the two color channels onto two separate cameras. The
general consensus was that you could "buy your way out of the problem"
by buying a thick, ultraflat dichroic.
I am trying a similar approach on a setup in our lab but, due to space
constraints that are not easily overcome, have tried first placing the
channel-splitting dichroic in the image space after the final tube lens
instead of in the infinity space between the objective and tube lens.
Even with a 3 mm thick ultraflat dichroic, I see moderately bad
astigmatism in the reflected channel. I have ruled out other possible
sources of the astigmatism, like an additional dichroic for the
excitation light and filters that already lie in the infinity space
between the objective and tube lens.
My question is: is it even worth trying to eliminate the astigmatism in
the reflected path of a split-channel setup if the dichroic does not lie
in the infinity space? Or is it pretty much always a bad idea to put the
splitter dichroic in the image space?
My current suspicion is that the tolerances in the alignment have to be
very, very tight to avoid astigmatism with the splitter in the image space.
Thanks for your feedback.
Kyle
--
Kyle M. Douglass, PhD
Post-doctoral researcher
The Laboratory of Experimental Biophysics
EPFL, Lausanne, Switzerland
http://kmdouglass.github.io
http://leb.epfl.ch
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