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Have you considered using single molecules as a fluorescence intensity
standard?  The real advantage is that they either emit, or are bleached out.
If you pick out a molecule that is bright and non-blinking, you might be
able to do a statistical study over a number of molecules to compare
brightness (eg. counts/second) at a known laser power.  The trick of course
is to locate just one molecule; this is most easily accomplished by spin
coating a very dilute (picomolar range) solution on a cover slip, so that
the mean distance between molecules is several microns.  This gets around
the issue of measuring the concentration accurately, or worrying about
whether the sample is bleached or not.  It is also a quite good test of your
entire optical system.  It is often helpful to spin coat a thin barrier
layer such as PMMA to reduce bleaching on top of the molecules.

Gary Holtom
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Pacific Northwest National Laboratory
phone 509 376 5331