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You really need to look at the whole Z stack to determine the problem. Would you put the TIF stack on Dropbox along with the capture metadata?


Eric Marino
Senior Imaging Specialist
Program in Cellular and Molecular Medicine
Boston Children's Hospital
200 Longwood Ave
WAB Room 133D
Boston, MA 02115
Lab: 617 713-8885
Cell: 617 913-9647
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On Sep 13, 2013, at 2:49 PM, Serkan Berk <[log in to unmask]<mailto:[log in to unmask]>> wrote:

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Hello,

I am observing some sort of an aberration (looks like coma) when imaging
sub-resolution sized fluorescent beads under epifluorescence. I am using a
Zeiss Axio Observer Z1 microscope with a Zeiss C-Apochromat 63x Water Obj.
NA=1.2.

I match the collar correction to the coverslip thickness but the airy disks
are not complete. I uploaded one of the images here
(https://www.dropbox.com/s/kst4plbrqmv29hn/100nm%20100x%201.0s.tif).

Image was taken with a sCMOS Camera (Hamamatsu Orca Flash v4.0) attached to
the side-port of the microscope. Particles are excited with SpectraX light
engine from Lumencor through a GFP filter set from Semrock.

Has anyone come across such problem? Let me know if you have any ideas about
what might be the possible causes?

Thanks
Serkan Berk

University of Minnesota
116 Church St. SE
Minneapolis, MN 55414