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List,

I am having some difficulty understanding exactly how "offset" as used by
OpenLab talking to a Hamamatsu ORCA is being applied.
My initial understanding of how this sort of function was applied was that a
fixed amount of current coming off the chip would be deducted from every
measurement, "setting black to a given value".
In OpenLab it is expressed as a percentage, but a percentage of what? The
range of intensities in a given captured frame? Or a percentage of all 12
bits the camera can produce?  In a few quick snapshots with transmitted
light I have failed to convince myself that it is working in a linear
fashion.

This has all come up because a user imaging YFP has reported that a 5%
offset (as opposed to 0) is making all the difference in detecting signal
over background, eliminating the need for histogram stretches.
I am concerned that a blanket hardware offset at collection (as opposed to a
calibrated pixel by pixel table) may be skewing (perverting?) the data when
we are working in the bottom end of the camera sensitivity.

Thanks for your thoughts,
Marc

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Marc D Green
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