Posted by
Andrew York on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Signal-bleaching-and-toxicity-vs-exposure-time-for-constant-total-light-dosage-tp7580528.html
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Simple question:
I take two 3D volumetric images of a cell, (widefield, 488nm laser
excitation, GFP-tagged).
In both cases, I take 10 slices over 1 second.
In case 1, I use 100 ms exposures, with a laser intensity of 1 unit.
In case 2, I use 50 ms exposures, with a laser intensity of 2 units. I turn
the laser off for 50 ms between exposures.
Which case bleaches the cell more? Which case is more phototoxic? Which
case gives the most signal? Surely someone has studied this, and the answer
is well known. I assume there's some linear range where total dose is all
that matters, and some non-linear range where this behavior breaks down.
Motivation: my users always want to turn down the laser and crank up the
exposure time, hoping this will be gentler to the cells or give lower
background or higher signal. I usually give my opinion that it won't make
much difference, but I don't have a solid reference to point them to for a
solid answer.