I would like to use Cellprofiler’s provided pipeline for the Comet Assay to analyse my slides.
But there are 3 things that I am unsure about or don’t seem to work properly.
In some situations the tails are relatively shorts, is there any way to be sure that the MajorAxisLength (or maximumRadius) actually is the length of the tail instead of the width?
Especially with longer tails Cellprofiler has a tendency to think the head is in the tail area or can’t find it at all. I assume it might help if I adjust the diameter for identifying the head but I am unsure which adjustments would help here.
I noticed that the program also counts the halo around the head as the tail. Since some of the data I have shows the MajorAxislength of the tails as long (or even longer) than the whole comet is this also used in the MajorAxislength calculations? (meaning I would get a more accurate tail length by substracting head length from the total comet).
I hope you can clarify these things for me or have suggestions how I can personalize the program to suit my experiment. I have attached an example what the maskobject images look like.
Thanks in advance!
(Images were made with 5 times magnification but the 10 times magnification images have similar object identification problems)