Line artifacts in confocal imaging

Hello all! I had the following issue with confocal imaging using Zeiss LSM 880. I notice horizontal lines on the objects, and they are more obvious on the edge of the objects. Can someone shed some light on what might cause it? Thank you.


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Might you have bidirectional scanning enabled?

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Thanks for the reply. That’s my first guess, but it’s uni directional scan.

I’m all out of guesses :joy:

I suppose it could be physical movement/vibration?

May be the scanners are going bad. What zoom and speed is this? Is this the collected image or the Live image?

Thanks for your help anyway.

It’s a collected image at 63x and 1.3x zoom. The speed is somewhere in the middle. Thanks for the input.

I would see if it persists after a system reboot. If it does, you might call your service tech to perform a scanner calibration.

As Talley mentioned, also check that your air table (vibration isolation table) is floating and nothing has been moved such that the top of the air table now has direct connection to the wall/floor/neighboring desk.

Good luck!

Thanks again. I agree the vibration could be the source. On the other hand, the artifacts are so regular, reminding of a bad AFM scanning pattern. This could be related to a bad scanner. I will talk to our FSE as well. Thank you both.

It’s hard to tell from such a low resolution image - but does this pattern disappear when the detectors are not saturated? These patterns can emerge when detectors are overexposed - more of a problem on ChS than Ch1 or Ch2 (but still does happen).

Otherwise I would test to see if you see this pattern on all channels with all laser lines - only then would I blame the scanners.

Edit: if it is a scanner issue you will generally see tearing across the whole image - not just where the brightest signal was.

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