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Gain is definitely confusing.
On some detectors used for point scanning, gain does increase sensitivity in a manner that the additional noise is ok. On other detectors, gain essentially increases noise. Different microscope software also have different ranges for gain. I have to adjust when I move from a Zeiss LSM with PMTs to one with GaAsP detectors to our older Leica SP8 to the Stellaris, and the Olympus systems gain appears to be all noise whereas voltage is signal. And on the new Stellaris, we have different detectors; at least two of them gain is pure noise after a narrow range where it increases signal, and after this point, accumulation is the preferred route to get better images. (Teaching people this is difficult.) Then there's the issue of gain in EMCCDs vs the other cameras we have.
I've quantified some of these, but mostly is just something I've noticed.
No, gain is not a one size fits all.
But there is one constant: users will saturate images to boost low signal areas instead of imaging correctly.
Cheers-
Michael Cammer, Sr Research Scientist, DART Microscopy Laboratory
NYU Langone Health, 540 First Avenue, office MSB 06 57, main lab Smilow C-17, New York, NY 10016
Office: 646-501-0567 Cell (DO NOT TEXT): 914-309-3270 [log in to unmask]
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-----Original Message-----
From: Confocal Microscopy List <[log in to unmask]> On Behalf Of Jeremy Adler
Sent: Thursday, April 27, 2023 5:41 AM
To: [log in to unmask]
Subject: PMT gain - what actually changes
[EXTERNAL]
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When the gain of a PMT is increased the output signal and therefore image changes - the point.
I assume there are different causes (i) the signal from individual photons is increased , (ii) more photons are detected and (iii) noise increases.
Detecting more photons - is this predominantly increasing the detection of lower energy photons?
If this is the case, the nice graphs of wavelength and QE will vary with the PMT's gain - so under which conditions are the graphs provided by microscope manufacturers generated.
Thoughts.
Jeremy
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B i o V i s P l a t f o r m of Uppsala University
Light & EM microscopy / FlowCytometry & Cell Sorting / Image Analysis ===============================================
Jeremy Adler PhD - Senior research engineer
Light, Confocal Microscopy, Image Analysis
E-mail: [log in to unmask]<mailto:[log in to unmask]>
070-1679349
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