Did you forget to ask your question?
Sorry some glitch I suppose. Yes I am trying to enumerate the cell count from ROI_1_B file while the nucleus is in ROI-1_G file. I was using the %positive pipeline. I am not sure if this would be the best way or I can get away only with ROI_1_G file. I am only interested in cell count from ROI_1_G file. I am new user so still trying to wrap my heard around this and image analysis.
I am not sure using only ROI_1_B would give correct results, since signal in some of the nuclei space is not even (I mean in the hole kind of region), so its better to use both the images. You could get the larger regions from the ROI_1_B i.e. by creating the mask (by thresholding) or selecting ROI. Use the ROI_I_G to threshold the nuclei & apply the mask or ROI to count inside the regions (i.e. the larger region in the ROI_1_B).