How to Analyse Particles PER Cell

Hello,

I would like to count the number (and preferably also the size) of particles per cell, in a given field of cells. I have added two example images which represent the two channels of my images, of which one shows the nuclei (BLUE) and the other shows the particles (GREEN).

Up until now I have made a macro that analyses the particles in the entire field pretty well. I can subsequently make an estimation of the number of particles PER cell by dividing the total particles by the number of nuclei:

macro “AnalyseParticles [7]” {

imageTitle=getTitle();
run("Split Channels");
selectWindow("C2-"+imageTitle); 
run("Duplicate...", " ");
run("Gaussian Blur...", "sigma=1");	
run("adaptiveThr ", "using=Mean from=20 then=-80");
run("Watershed Irregular Features", "erosion=1 convexity_threshold=0 separator_size=0-Infinity");
run("Make Binary");
run("Analyze Particles...", "size=0.10-1 display exclude clear add");
run("Summarize");

}

macro “AnalyseNuclei [8]” {

run("Gaussian Blur...", "sigma=2");
run("Auto Threshold", "method=RenyiEntropy white");
run("Analyze Particles...", "size=40-Infinity display clear add");
run("Summarize");

However, I would like to segment each cell based on the channel 1 (Nuclei) and then analyse the particles PER cell. I tried segmenting the cells using the Find Maxima function in the Nuclei channel, which worked reasonably well, yet I was unable to analyse particles in the segmented areas. Hence, I am desperately looking for your help!

Thanks in advance,

Maarten

Hi @Maartench,

since you have nuclei and puncta already segmented you can have a look at the “Speckle Inspector” from the Biovoxxel Toolbox.
When you run an update on Fiji you can add the BioVoxxel update site and this gives you the tool mentioned to analyze puncta per cell/nucleus starting with the two channels segmented as a binary image (no need to run an individual Analyze Particles… command in your macro. The function is then also macro recordable.

hope that helps you further.