RGB colocalization in ImageJ

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oojeyboojey
oojeyboojey's picture
RGB colocalization in ImageJ

Hi,

I've been doing some multiphoton imaging of nanoparticles at a variety of wavelengths.  I now have separate images of the same samples at three wavelengths and want to combine the images to show only the regions of colocalization. 

However, I need the pixel contributions from each of the three channels to only be displayed if they're at a specified ratio to each other, i.e. each pixel in channel A that is 2x the intensity of the corresponding pixel in channel B is selected and all others are discarded etc.

I've seen an interesting plugin:

grove.ufl.edu/~ksamn2/plugins.html

But I don't really know how to express the ratios as percentages.  For instance, if I only want to display the green channel's pixels if they're 2x the intensity of the corresponding pixels in the red channel, how do I do that?

Does anyone here know how to help?

oojeyboojey
oojeyboojey's picture
OK I figured out the ratios

OK I figured out the ratios as a percentage thing (i.e. 1:1 ratio is expressed as 50%, 1:2 ratio is expressed as 33.3% etc.) but I need to know if this plugin is screening out colocalized pixels which don't match this ratio, or if it's adjusting their intensities so they're at the specified ratio.  If it's screening them out, what's the tolerance?

I guess I could try to figure this one out for myself by playing with it for a couple of hours, but if anyone here happens to know, please save me time I could be spending writing my thesis!