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Am 13.03.2015 14:21 schrieb "Eric Shelden" <[log in to unmask]>:
>40X water immersion 1.1NA Corr lens
> should have given better resolution and better brightness
> than our 20X 0.7 Corr multi-immersion lens.
> it provided only slightly better resolution and
> prequired higher gain settings to achieve saturation with the same laser
> power settings. I was wondering if there was a reasonable explanation

Eric,

you can measure the angle of light emitted from small but bright objects in
your specimen or of beads. xz scan through a couple of beads or objects.
The sinus of the full angle multiplied with the refractive index of your
media gives the NA of the detection system. To make sure to achieve full
resolution you also need to make sure to overfill the back aperture of the
objective. There is a beam expander that should be set to the largest
number.
Make sure to also adapt the magnification to integrate photons from the
same area. Without that you would expect a lower intensity for the 40x,
because you integrate from a smaller area.

Hope it helps,  Jens

Visiting Scientist @ Center for Technological Development in Health (CDTS),
Oswaldo Cruz Foundation (Fiocruz), Ministry of Health, Rio de Janeiro,
Brazil