Counting 3D immunofluorescent particle

[ergefan]

Hi All,

I wonder if anyone could give me some advice on counting fluorescent particles in a stack.

I am now using 3D object counter to count the number of particles and compare between control and experimental group. Images from experimental group are more fluorescent and the particles look more clumpy. The problem is that when I set the threshold at low value, the control would have more particles than experimental group because the particles in experimental group with higher fluoresence would clump together and be seen as one; when threshold is high, the experimental group would have more particles than the control because the puncta with lower fluorescence would not be included for counting. My aim is to count every particle in the images (with information about particle fluoresence intensity and area).

Any idea how to solve this?

Thanks very much for helping.

Best wishes,
Kai.
Postgrad in The University of Sheffield.