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Date: | Wed, 25 Jul 2001 16:45:04 -0400 |
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This is an Interesting observation with a question for the confocal user
group
I was trying to do Huygens deconvolution ( Bitplane software) to increase
the resolution of some confocal images. In order to do this effectively you
are supposed to use 0.17 micron beads ( balls) or 0.5 micron beads( balls)
with high power objectives to get a reference particle for the software.
While doing this test to acquire the necessary reference images from the
0.17 u beads, they appeared as doublets( two beads) and the 0.5 micron
beads appeared egg shaped. What will this type of distortion observed on
beads do to the fluorescent images of cells and particles?
On closer inspection we notice the Wallistron prism, which is necessary
for interference contrast, was in the light path. I interpret these results
to mean that while doing interference contrast simultaneously with
fluorescence there will be a degradation of the fluorescence signal. Does
any one know by how much? Should we not use interference contrast when
acquiring high resolution fluorescent confocal images?
Although interference contrast is a useful signal, it should not degrade
the fluorescence signal if ones wants optimum resolution from a specimen. I
would like to have some input on this observation and any suggestions or
recomendations on the simultaneous use of both signals. Thanks
Bob
Robert M. Zucker, PhD
U.S. Environmental Protection Agency
MD 72
National Health and Environmental Effects Research Laboratory
Research Triangle Park, North Carolina, 27711
Tel: 919-541-1585; fax 919-541-4017
e-mail: [log in to unmask]
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