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 `Histogram2D`

and `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?

/cc @etadobson, @chalkie666