Dear colocalization and signal-processing experts,
for one of my projects I need to determine the crosstalk/bleedthrough between two fluorescent channels. The experimental setup necessitates that we acquire at suboptimal conditions where one channel has a significant bleedthrough to the other channel (Ch1 -> Ch2) but not the inverse (Ch2 -> Ch1).
To do some simple channel unmixing, we’d like to measure the intensity correlation of the two channels in images that contain only one of the fluorophores (Ch1). I know that Coloc2 does this calculation by linear regression on a 2D histogram, but I’m looking for an easy way to determine a possible non-linear relationship between the two channel intensities (since I’m not sure that we are working in the linear range of our dual-cam setup).
I had a quick look at the
AutoThresholdRegression classes of Coloc2 to see if I could re-use some of their (linear) regression logic, but the code seems to be pretty baked in to the specific use case of colocalization measurement.
I also checked out the Spectral Unmixing plugin, but it does linear unmixing, where I would like to be able to define a non-linear function.
Is there any other library available for non-linear fitting of a 2D intensity histogram? Anything available in ImageJ-Ops already?