BioFormat Bug with stitched czi files

Hi,

i am using bio-format to import multiwell stitched image in a czi file.

I have this bug where the image is not read correctly by bio-format (see picture). The image is fine when opened in ZEN.

bug

I can save the image in tiff, but for the workflow to be easy and fast i would prefer to be able to open the czi file.

Any idea ?

Thanks in advance!
Rémy

1 Like

Hi @r.flores, if you are able to upload the file to https://www.openmicroscopy.org/qa2/qa/upload/ for testing that would be a great help.

Are any of the dimensions for the various series incorrect?
Does any of the other metadata differ from what you might expect?

Hi,

i have uploaded a file (id 27461).

As far as i’m aware the dimensions are correct. It worth noting that, when opened with ZEN, the origin for the image is not (0,0), but the position on the stage, (13971,61) for the second well.

For my script i’m reading metadata, but the original one, to find the well names (A1, A2, …), so i dont know if there an issue with the OME metadata.

Thank you for your help,
Rémy

@r.flores I encountered something similar with stitched images and I am not sure Bioformats is the place where the bug happens.

In our case we noted that the X,Y,Z position of each tile reported by Bioformat was very wrong. Could you post here the positions reported for each tile in your case?

The file contains several already fused mosaïc images corresponding to different well. The X,Y position i read with bioformat is the same as the top left pixel positon on ZEN.

So i suppose X,Y position correspond to the position of each image (so well) on the stage, relative to the first wich is (0,0).

Can you run this script in ZEN on your CZI image (in case you have the module for that). If not, can you upload it somewhere where I have access?

Sebi (from ZEISS)

@sebi06 i don’t have the option.

So i have uploaded for you the original file and the stitched one, which i have an issue to open with bioformat with.

https://filesender.renater.fr/?s=download&token=b9cf4337-fe8d-cb1e-241a-c5439557a628

Thanks,
Rémy

So I attached the two plane tables for you. How did you do the stitching in ZEN? Did you fuse tiles for the stitching?

Not stitched:

OCR-LySM-CPS_PlaneTable.csv (29.9 KB)

Stiched:

OCR-LySM-CPS-stitched_PlaneTable.csv (1.7 KB)

Maybe this helps to find the problem. But when I try to open the un-stichted and the stiched CZI with the latest Fiji it looks OK to me. (I only opened the 1st scene using the hight resolution from the image pyramid)

My Fiji is using BioFormats 6.0.1

@sebi06 yes i fused the tiles for the stitched images.

The first image is always fine with Fiji and Bioformat, the bug appears afterwards.

My czi file doesn’t contain an image pyramid, but different scene.

Hi,

hmm… If I open other scenes from the file you sent me, it also works fine in Fiji. But why does your CZI does not contain a pyramid. Did you switch this of on purpose?

But I agree, from your screenshot I see your are right. So maybe BioFormats cannot deal with CZI images that do not have a pyramid?

OK, so it seems that BioFormats 6.0.1 cannot read stitched CZI images correctly, where you fused the tiles.

I did a further test. IF you stitch the images in ZEN and do NOT fuse the tiles, that it works fine in Fiji wen opening the image via BioFormats 6.0.1. But the the CZI still has a pyramid

So the issue is: BioFormats 6.0.1 cannot open stitched CZI images where the individual tiles where fused after the stitching.

1 Like

@sebi06 ok thank you I’ll try without the fusing. I’m actually not sure if the file with the fused images i send you as a pyramid or not.

You can see this when opening the CZI via BioFormats and look for the image dimensions inside the series dialog. Check also this post:

2 Likes

@sebi06 so i have tried to stitch without fusing. But when i want to open then with bioformat if i dont check the option stitch tile it only open one tile of the scene and if i check it want to stitch all the tiles from all scene, which is a 150528 by 156672 image !!!

Thank you all for the detail in this thread, it’s very helpful. We now have the OCR-LySM-CPS-stitched.czi file uploaded here safely at OME. I am reminded of the older thread at https://lists.openmicroscopy.org.uk/pipermail/ome-users/2017-August/006631.html – maybe a related issue?