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There ws a paper a few years back which took a contrary view.  I
guess it depends on how fanatical you are going to be.  As to
deconvolution, if you are going to recover your psf by deconvolving
from an image it might be better to use a larger object which can
be more easily characterised - a grid for example.  But 100nm beads
give you an easy, direct visualization of your psf, and the new 60nm
beads we've bought seem to be about as bright so at any rate they
can't be worse!

                                                        Guy

>Guy Cox wrote:
>
>> Some points - if you want to measure a true PSF of a high NA lens
>> 100nm beads are the maximum you should consider, and 60nm which are
>> now easily available would be better.  Oversample as much as you
>
>I'd dare say 100nm is plenty small enough. Since the PSF will only be
>enlarged due to the *convolution* with a 100nm bead the error with a
>200nm in-plane resolution is ~10%. Given a 500nm out-of-plane resolution
>the error is down to ~2%. An if you are really fanatic about recovering
>the actual values you can deconvolve the bead (100nm sphere) from the
>recorded PSF.
>
>
>   Christian Soeller
>
>--
>Christian Soeller PhD           Dept. of Physiology
>University of Auckland          Auckland, New Zealand


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