Binary-Convert to Mask does not work

Sample image and/or code


This is a mouse brain implanted with tumor, the immune cells were engineered with Green fluorescence protein or red fluorescence protein, and we stained the nuclei with blue. So there are green, red, and blue in the slice. We need to quantify the green cells and red cells. The histology images were acquired using a machine named NanoZoomer, the raw data are. NDP files, we review the images with NDP viewer, calculate the area of the tumor, make a screen shot, then quantify the cells from the screenshots with FIJI.

Analysis goals

Quantify Green and red cells


We draw the outline of the tumor in FIJI, then Edit-Clear outside-Subtract Background with 4.0 Pixels-Separate channels to red, green, and blue. For the Green or Red separated images, we do Pluggins-Filter-Otsu Thresholding, then we go to Process-Binary-Convert to Mask. Usually the background will become white when we run Convert to Mask, but with this IMAC (it is a new computer, we installed FIJI, Java, Otsu thresholding pluggin) it doesnot work. I ran the same steps, it works on my Macpro book.
Thank you for the help in advance!

When you say does not work, do you mean it is backwards or you get an error? I believe macs and PCs can have the black/white settings reversed, and you may need to set your background to black.

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Thank you soooo much, I unchecked the “Black background” just now, and it worked!
I really appreciate!

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Good afternoon, I ran into another problem with FIJI, after running Binary-Convert to mask, I need to run Binary-Watershed, there is no change afterwards, could you please help me?
Thank you!

Afraid the only thing I can think of, based on your previous issue, is that you might not have the correct black particles on a white background?

If you have it set up like that (no invert image set), then you may want to start a new topic and provide a sample image.

I already have a white background after running Convert to Mask.
OK, I will start a new topic.
Thank you!