My question pertaining to the DAB staining in cytoplasm of human oral squamous cell carcinoma tissue.
When quantifying the epithelial cancer cells do we have to crop remove the stromal tissue? because stromal positivity is also seen in the sections.
Any separate plugin in image j or any other open source software to quantify stromal positivity?
kindly share your valuable suggestion.
Dear Mohamed Nassar,
When quantifying epithelial cytoplasmic DAB staining using the IHC PROFILER plugin in ImageJ, it is essential to remove or crop out the stromal tissue, especially if stromal positivity is observed in the sections. The stromal staining is considered background when epithelial markers are used in immunohistochemistry, and it should not be included in the analysis. Only the epithelial staining should be quantified.
To specifically quantify stromal positivity, you can use the Region of Interest (ROI) Manager in ImageJ to manually select and analyze stromal areas. Alternatively, you might find QuPath to be a valuable tool. QuPath is an open-source software designed for digital pathology that offers advanced features for tissue segmentation and quantification, allowing you to differentiate between epithelial and stromal regions more effectively.
Dear Petro V. Kuzyk Thank you sir for your response. when I crop out the stroma, that area remains dark. During quantification, the histogram shows a peak in the darkest intensity zone. Alternatively, if i invert the dark areas into white areas, there is a spike in the lightest zone in the histogram eg. 255 pixel intensity region. have you come across such situation?
Dear Mohamed Nassar,
Thank you. When cropping out the stromal tissue, the dark area left behind can indeed affect the histogram analysis, leading to peaks in the darkest intensity zone. This issue arises because the cropped area is interpreted as part of the analysis, skewing the results. To address this problem, you can try the following options:
Dear Petro V Kuzyk, may I suggest you share a demo video of the way mentioned here? Thanks a lot.
Dear Mohamed Nassar,
Thank you for your message. I suggest sharing a demo video to illustrate the method mentioned for analyzing DAB staining in epithelial cancer cells while excluding stromal tissue.
Best regards, Petro V. Kuzyk
Dear Petro V. Kuzyk, find attached the video to know the way i crop removed and inverted background color instead of black to white and then quantified the IHC DAB staining.
you may please send me demo video of the ways you suggested above. particularly with the ROI manager method.
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