> >I think that when you play with the contrast of your image during
> acquisition, you change the PMT gain. Is that true?
> >I am also wondering what is affected when you play with the brightness?
> >Thank you very much for your help.
>
> In my experience it is true that changing the "contrast" of the image
> aquisition changes the gain of the PMT. You should also note that it does
> not have a linear effect on the image. Usually the "brightness" control is
> just an DC offset that adjusts the black level of the image so that very dim
I guess this probably depends on what system you're using.
In the Zeiss LSM 410 (I don't know about LSM 310 and/or 510) the PMT
voltage AND the amplifier gain are changed simultaneously with the
contrast command. The sensitivity (an internal variable) will tell you the
individual settings of the PMT voltage and gain. However, you can
separately control the PMT voltage and the amplifier gain (you need to
write a macro for this) exactly the same as with a flow cytometer, for
example. Obviously, image intensity will vary non-linearly with PMT
voltage. However, it varies linearly with amplifier gain. It seems to me
that this feature hasn't been exploited extensively by LSM 410 users.
In practice, it's almost equivalent to a larger dynamic range. It seems to
me that controlling gain and PMT voltage separately also gives better
control over noise.
As you said, brightness controls an offset current. I presume that the 0
value for that current has to be determined for each system. In our
microscope is 9555 (Zeiss UK told me that it's usually around that value).
Hope it helps

Carlos