Thanks for all the additional information. I believe I have enough information to help me to generate some expression heat maps from the ISH images now.
What I’ve decided to do is to take the ISH color information, and apply a transform in color space so that the “purplish” colours end up being aligned with one of the color space axes (it happens to be the one that was originally ‘blue’). I then make an elliptical tube around this axis, and all color pixels whose (transformed) color lands them inside the tube get a non-zero “expression” value taken from the projection of the point onto the axis. If the pixel is on this axis, but it nonetheless too ‘light’ then it gets expression=0 assigned to it.
It would still be great if the precise algorithm that was applied to turn ISH images into expression maps was published at some time. Is the code that was used open source or available by personal communication?