make a recommendation practical of how to measure intensities in spots

[tischi]

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[tischi]

1. 8-bit is enough! A colleague of mine did once a calculation that the information content in a confocal (in terms of actually collected photons) is typically only worth 5-bit.

2. you should do a local background subtraction (https://github.com/tischi/imagej-courses/blob/master/imagej-practical.md#local-background-subtraction) and then measure the integrated intensity of each in a generous(*) region of interest that encompasses all pixels that belong to the respective dot.

• due to the local background subtraction pixels around the dot have zero intensity so it does not matter if you include them in the integrated intensity, thus your ROI can (and should) be generous.