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Kathryn Spencer wrote:
> 	I was also impressed with the STED microscope at SFN. Great
> improvement in resolution. However, as Martin said, it only works with
> two dyes now (although someone brought a sample with them to the demo,
> and found that their dye also worked...now there are three), and only
> one dye at a time. 

...Just to make sure that this point is clear--it can deal with more 
than one dye, but the dyes that can be used are all far-red (i.e. 
emission between about 650 and 700 nm).

> The system is only good for fixed samples...it didn't
> appear to even be useful for cell surface labeling of live cells.
> Because of the requirement of having two lasers at different wavelengths
> perfectly aligned, penetration depth was limited to 15-20 microns. I'm
> still sending them some samples from our labs, but I too am waiting for
> the "second-generation" STED before I send my PO.

When I spoke to the Leica people, they said (as best I recollect) that 
development of multi-color STED depended on development of the 
appropriate fluorophores and that there were no hardware improvements 
that were in the immediate pipeline.  That's a bit surprising since (as 
was pointed out to me off-line) Gerald Donnert in Stefan Hell's group 
has already published on multicolor STED (Biophys. J. 2007 92: L67-69L) 
and hints that improvements in lasers and related hardware should make 
implementation easier in the near future.  --In that paper they also 
mention the depletion laser for the far-red fluorphore as not being 
optimal...which again seems odd since far-red STED is what Leica is 
selling.

My aged memory has probably turned the story around 180 degrees, though, 
and I should probably invite the reps from Leica chime in and set the 
story straight!  Maybe they could also comment on how easy (or 
difficult) it is to keep the microscope aligned and the related issue of 
improving depth penetration.

Martin Wessendorf

-- 
Martin Wessendorf, Ph.D.                   office: (612) 626-0145
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