Posted by
Daniel White-2 on
URL: http://confocal-microscopy-list.275.s1.nabble.com/why-does-high-NA-excitation-illumination-give-better-resolution-in-fluorescence-microscopy-tp7581519.html
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Hi again all,
So from the informative answers so far...
it seems that the idea of filling the back focal plane of the objective
(acting as the condenser lens) in epi fluorescence, both widefield and
confocal, is more about getting lots of light into the sample in order to
get enough signal to then be able to see the resolution that is there,
since effective resolution is limited by signal to nose (sorry, noise)
In other words, is there enough light going in (and consequently coming
back out) that we can see the Rayleigh criterion dip between the two peaks
of the object images, despite any noise that's present?
Is this following statement then true?:
In the case of infinite signal to noise, where contrast is optimal and not
limited by noise, where illumination power is close to saturation of the
fluorescence excited state, exposure time is long enough, etc....
the effective NA of illumination (back aperture filling) has no effect on
achievable lateral or axial resolution - only signal:noise does.
Are there cases where this is false? Anisotropy?
cheers
Dan