Posted by
James Pawley on
URL: http://confocal-microscopy-list.275.s1.nabble.com/DIC-or-Phase-contrast-for-image-morphometry-mesasurements-tp7068286p7070645.html
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>I did a lot of yeast imaging with DIC in grad. school and it was a
>pain for morphometry because of the "shadow" effect of DIC; one side
>of the cells was bright while the other was dark and that was
>problematic for thresholding. Now I'd use phase, or even brightfield
>or darkfield. From your question, though, it sounds like getting the
>cell outline is no problem for you with DIC and you're just wondering
>about accuracy, which I don't know.
>
>-Esteban
>
>
>On Tue, Dec 6, 2011 at 12:23 PM, Vasseur Monique
><
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>> Dear All,
>>
>> I wonder what is better for yeast morphometry shape and size image analysis:
>> should we better use phase contrast images or DIC images? Is one more
>> accurate for measures? )Which threshold will be more precise: the
>>one of DIC
>> shear or the one of phase contrast halo?) Do some of you have experience on
>> this question? Any input is welcome Thanks a lot!
>> Monique
Dear Monique,
As everyone is chipping in, have you thought about using darkfield?
You would need a good darkfield condenser and they can be a bother to
set up (though a lot cheaper than DIC and not too bad setting up on
such a thin specimen.) and an objective with an NA about 1.15 (or
larger but with an iris diaphragm. Of course, you would lose a little
on the resolution because of the lower NA (but not that much if your
specimen is in water as you don't get many rays propagating past NA
1.25 anyway because of reflection losses.
What you do gain is a very simple image (objects having a different
RI from their surroundings look white on a black background). And, on
such an image, it might be much easier to come up with an accurate
and reproducible measurement scheme.
It would probably not be hard to get your microscope salesman to come
around and set up a system for you to see how well it works.
Jim Pawley
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"If it ain't diffraction, it must be statistics." Anon.