Dear experts, I am a student from China. Recently, when I used the light sheet7 (Zeiss) light microscope to image the pancreatic islets, there was blurring. However, it was very clear during the two-dimensional scanning. Moreover, when I used imaris to reconstruct the three-dimensional image, a strange image appeared. How can I set the shooting parameters to make the imaging clearer? What other commonly used analysis software can be used in the later 3D image processing besides imaris software?
I would recommend talking to the Zeiss rep, but it is difficult to know what might be happening without some understanding of what the blurring looks like, what kind of 2 dimensional scanning was done (different instrument? preview mode on the lightsheet?), what the sample/sample holder is, and what “strange image” means more exactly.
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If the blurring is due to convolution with a point spread function it can be partially corrected with deconvolution. There is a commercial product called Huygen’s (I have no affiliation with the company) that can be used for deconvolution.
In addition Imaris has a plugin called Clear View which may also be used for deconvolution, though I am not sure which algorithm they implement.
There are also many open source plugins for ImageJ and for Python/Napari which will give just as good results, though can be harder to use. Many people have had success using the open source plugins by opening up a question on the sister ImageSC forum
Without seeing your images I can’t say for sure if Deconvolution will help. The key to deconvolution is to measure a high quality Point Spread Function, so most of the work will be in figuring out how to do that and may involve collecting bead images.
Brian
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When you say ‘reconstruct’ which algorithm from Imaris did you use? Was it the deconvolution plugin? Sometimes people simply use ‘reconstruct’ to mean taking 2D slices of data and generating a 3D rendering. However if you did apply Deconvolution or another algorithm first, you may have used an incorrect PSF or incorrect settings. People will need more details to offer useful advice.