Re: 2P standard

Posted by George McNamara on
URL: http://confocal-microscopy-list.275.s1.nabble.com/2P-standard-tp5923717p5929249.html

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Hi Carl,

Instead of using a bleachable fluorescent target, use something that
makes great second harmonic generation (SHG). If you MP laser tunes from
690-1040 nm an SHG specimen will be usable from 395-520 nm (maybe you
can convince NIH's NCRR to give all of us S10 money to add an OPO or OPA
to our MP systems to extend the wavelength range).

Besides sugar crystals, starch granules, collagen fibers (see Rich
Cole's post), Scott Fraser's lab published a PNAS  paper in 2010 using
nanoparticles for multicolor SHG-immunofluorescence (antibody
conjugation procedure developed by a different Caltech lab and cited by
Fraser and colleagues). They used specific types of particles from    
http://www.nanoamor.com/products
Multicolor SHG worked because some types of nanoparticles perferentially
underwent SHG in specific excitation wavelength range, whereas others
had broad excitation range.

Enjoy,

George



On 1/14/2011 6:34 PM, Boswell, Carl A - (cboswell) wrote:

> Hi all,
> I'm looking for some trustworthy, permanent standard(s) to use in a MP system to monitor the condition of detectors and the system in general.   The goal is a fluorescent source that is immutable.  This could done either by having unrestricted replacement of bleached label with unbleached label (i.e. a solution), or a uniform and very large solid, so that it is unlikely that any one tiny volume will be imaged repeatedly.   We've tried several iterations of an aqueous solution of label, but sealing the prep does not prevent eventual evaporation.  Plus, there seems to be some modification of FITC, for example,  in solution over time (a month) regardless of exposure to light.  I'm not sure I can trust the consistency of colored plastic slides, since they are not manufactured with this role in mind.  One possibility is uranyl glass slides, but Corning says they don't make these any more.  Does anyone know a source?  We're also considering quantum dots in something like immersion oil , in a sealed chamber but I don't have any experience with these labels in a hydrophobic environment.
>
> Thanks for your insights.
> C
>
>
> Carl A. Boswell, Ph.D.
> Molecular and Cellular Biology
> Univ. of Arizona
> 520-954-7053
> FAX [phone]
>
>
>