Sorry @etadobson.

I was a little bit short of time yesterday.

I edited my post above. It’s rather a conclusion than question.

Here comes the real question: I selected a ROI in the image I posted awhile ago (see above),

removed the background and split the channels into R, G, and B.

Still I’m getting a high correlation, where I’m visually

not seeing any (the amount of red and green pixels in the image above should be quantified):

```
!!! WARNINGS !!!
Warning! y-intercept far from zero - The ratio of the y-intercept of the auto threshold regression line to the mean value of Channel 2 is high. This means the y-intercept is far from zero, implying a significant positive or negative zero offset in the image data intensities. Maybe you should use a ROI. Maybe do a background subtraction in both channels. Make sure you didn't clip off the low intensities to zero. This might not affect Pearson's correlation values very much, but might harm other results.
Warning! Threshold of ch. 2 too high - Too few pixels are taken into account for above-threshold calculations. The threshold is above the channel's mean.
RESULTS:
Coloc_Test
% zero-zero pixels, 4.97
% saturated ch1 pixels, 0.00
% saturated ch2 pixels, 0.26
Channel 1 Max, 255.000
Channel 2 Max, 255.000
Channel 1 Min, 0.000
Channel 2 Min, 0.000
Channel 1 Mean, 1.724
Channel 2 Mean, 14.599
Channel 1 Integrated (Sum) Intensity, 10233194.000
Channel 2 Integrated (Sum) Intensity, 86631428.000
Mask Type Used, none
Mask ID Used, 196894066
m (slope), -278.89
b (y-intercept), 495.54
b to y-mean ratio, 33.94
Ch1 Max Threshold, 1.00
Ch2 Max Threshold, 255.00
Threshold regression, Costes
Pearson's R value (no threshold), -0.03
Pearson's R value (below threshold), -0.03
Pearson's R value (above threshold), -0.04
Li's ICQ value, 0.090
Spearman's rank correlation value, -0.03011675
Spearman's correlation t-statistic, -73.3977
t-statistic degrees of freedom, 5934094.000
Manders' M1 (Above zero intensity of Ch2), 0.676
Manders' M2 (Above zero intensity of Ch1), 0.678
Manders' tM1 (Above autothreshold of Ch2), 0.001
Manders' tM2 (Above autothreshold of Ch1), 0.678
Kendall's Tau-b rank correlation value, -0.0211
Costes P-Value, 0.00
Costes Shuffled Mean, 0.00
Costes Shuffled Std.D., 0.00
Ratio of rand. Pearsons >= actual Pearsons value , 1.00
```

Hope somebody has a suggestion.

Best wishes,

Stephan