I’m trying to analyse my multiplex images using version Qupath 0.2.0-m8. I scan my images (.ndpi format). Using Qupath I select specific areas within my tissue and I send them to Image J using a downsample factor of 2. Then I align that images on Image J, so I end up having a 8bit stack image with z= number of markers I have (5 in my specific case, z=5) in a .tiff format. Then, still in Image J, I transform stack to images and I merge all of them assigning a different colour to every image and inverting LUT. Once I set up the brightness and contrast for every marker I save the merge image in OME.TIFF. The problem is visible at this point, when I open the image with Qupath to proceed with segmentation. I can see the colours are all inverted, background is the color I assigned to my marker of interest, and therefore the program is just analysing that background. Could you please help me with this? I have the feeling my problem is image processing related but I’m unable to discover where the problem is. BTW, the example image in 32bit works perfectly in Qupath 0.2.0-m8, so my understanding is Qupath hasn’t any problem.
Thanks a lot in advance for any help and feedback I could have