Currently I’m trying to measure the amount of nuclei( stained blue) in a population that are positive for BRDU (stained red). The problem I’m having is the intensities between each nuclei image. I’ve kept the exposure time the same when capturing them but some parts of the well have brighter background than others this is causing the pipeline I’ve created, to work within a narrow intensity band often rejecting images that are showing nuclei but are too weak in comparison to other brighter objects.
I’m hoping to find a way to normalise all the nuclei images background to each other, so that the brightest background images will be reduced and the pipeline will pick up all the nuclei in each image.
I’ve attached some examples of the difference between the backgrounds and the pipeline I’m using.
2brdu 17102013.cp (15.6 KB)
I’ve tried some rescaling and masking the image with some limited success.
Any help in improving the robustness of this pipeline would be greatly appreciated.