Re: 2P standard

Posted by Mark Cannell on
URL: http://confocal-microscopy-list.275.s1.nabble.com/2P-standard-tp5923717p5929332.html

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Since SHG is highly dependent on phase matching how do you ensure that  
the sample has exactly the same orientation and control the  
backscatter needed to test the PMTs?

Further to an earlier post,  my 4 year old plastic plastic slides look  
as good as new -but then I've looked after them and not exposed them  
to sunlight.

Cheers

On 17/01/2011, at 3:38 AM, George McNamara wrote:

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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 520-621-3709
>>
>>
>>