[OpenSPIM] focal part changing when I move beads

[Wiebke Jahr]

Hello Yan,

I'm guessing that you're facing two problems at the same time:

1) a tilted lightsheet that you need to align
2) a changing optical path length as you're moving the cuvette.

To be sure, could you please clarify:
Do you use the original open SPIM design with water-filled chamber and water dipping lens? The beads would be embedded in agarose, inserted into the chamber from the top and moved through the medium. In that case, changing optical path wouldn't be a problem.
Or do you use a modified design with an air lens, looking into the chamber? Do you then move the whole chamber (you're writing about a "cuvette") through the light sheet? Then the optical path would change as you move the chamber. As a result, the plane that your detection path focusses at would change. In combination with a tilted lightsheet, you'd get the feeling that the "in-focus" portion of the image translate.

To test for a tilted light sheet, just turn the bottom screw of the corner mirror (the one that's installed first in the original openSPIM instructions). If the light sheet is not tilted, it will just go out of focus over the whole field of view. If it's tilted, the in-focus part of the image translates, because the line where your tilted light sheet and the detection focal plane overlap shifts along the image.

Best, Wiebke

--
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Wiebke Jahr
PhD Student
Tel. +49 351 210-2768

Huisken Lab,
Max Planck Institute of Molecular Cell Biology and Genetics
Pfotenhauer Str. 108, 01307 Dresden
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----- Original Message -----
From: "胡衍" <huyan1020 at sjtu.edu.cn>
To: openspim at openspim.org
Sent: Friday, December 8, 2017 5:52:27 AM
Subject: [OpenSPIM] focal part changing when I move beads

Hello, everyone.

I used my openSPIM to take photo for beads, 
and found the focal part is always changing when I move the beads, as shown in the attachment.
The beads are in a cuvette.

Is there anyone knows the problem?
What can I do to solve this?

Best,
Yan
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