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Actually, that is a very good use of 2P and it works really well  
(shameless self citation follows)
See:


Cannell, M.B., Jacobs, M., Donaldson, P. & Soeller, C. (2004) Probing  
microscopic diffusion by 2-photon flash photolysis: Visualization of  
isotropic and anisotropic diffusion between lens fiber cells. Micr.  
Res. Tech. 63(1) 50-57

  Soeller, C., Jacobs, M.D., Jones, K.T., Ellis-Davies, G.C.R.,  
Donaldson, P.J. and Cannell, M.B. (2003)  Probing microscopic  
transport and release fluxes in cells by two-photon flash photolysis.  
J. Biomed. Opt. 8:418-427

Soeller, C. & Cannell, M.B. (2002) Estimation of the sarcoplasmic  
reticulum Ca2+ release flux underlying Ca2+ sparks. Biophys. J. 82.  
2396-2414.

Cheers


On 28/04/2011, at 12:48 PM, Rosemary White wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Dear all,
>
> Haven't followed this discussion closely, but would one advantage of  
> 2P lie
> in the ability to uncage caged compounds - fluorescent tracers, for  
> example,
> within a single cell?  We've used UV uncaging and while you get most
> uncaging in the target cell, you also get cones of uncaging above  
> and below
> the plane of focus.  Seems that 2P would overcome this problem.
>
> Rosemary White
>
> Dr Rosemary White
> CSIRO Plant Industry
> GPO Box 1600
> Canberra, ACT 2601
> Australia
>
> T 61 2 6246 5475
> F 61 2 6246 5334
> E [log in to unmask]
>
>
>
> On 28/04/11 10:10 AM, "Craig Brideau" <[log in to unmask]> wrote:
>
>> *****
>> To join, leave or search the confocal microscopy listserv, go to:
>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>> *****
>>
>> I second the previous lister's opinions.  At the end of the day MP  
>> is really
>> only good for thick tissue sections.  If you are looking at cell  
>> layers just
>> use a conventional 1P confocal.  The key advantage of 2P is  
>> penetration
>> depth, which is moot when you are looking at cells.  That said,  
>> John's
>> comments about shorter UV imaging are still valid; it's easier to  
>> get a NIR
>> Ti:Saph beam through conventional optics than a very UV beam.  This  
>> only
>> applies if you are using dyes that need very UV excitation of course.
>>
>> Craig
>>
>>
>>
>>
>> On Sun, Apr 24, 2011 at 6:29 AM, George McNamara
>> <[log in to unmask]>wrote:
>>
>>> *****
>>> To join, leave or search the confocal microscopy listserv, go to:
>>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>>> *****
>>>
>>> Hi David,
>>>
>>> The tuning range of Ti:Sapphire (ex. 690-1040 nm) used to be  
>>> problematic
>>> for exciting far red and infrared fluorophores. There were some  
>>> exceptions,
>>> such as Alexa Fluor 488m 568, 594, and 633 all exciting well  
>>> around 755 nm
>>> (Fig 5D of Dickinson et al 2003 J Biomed Optics 8: 329-338), but  
>>> since you
>>> are ok with cost and complexity, adding an OPO(s) and/or multiple  
>>> MP lasers,
>>> gives you full range (see
>>> http://www.coherent.com/products/?1570/Chameleon-OPO-Tuning-Range  
>>> for
>>> OPO).
>>>
>>> Depending on cell type and physiological needs, you may be able to  
>>> suppress
>>> phototoxicity by lowering O2 - cell culture at ~18% O2 is  
>>> something of an
>>> artifact for many cell types - by using catalase/glucose oxidase/ 
>>> glucose or
>>> Oxyrase (for the latter, www.oxyrase.com, see Waterman-Storer 1993  
>>> and
>>> Mikhailov 1995 Cell Motil Cytokseleton).
>>>
>>> Enjoy,
>>>
>>> George
>>>
>>>
>>> On 4/23/2011 6:03 PM, David Knecht charter wrote:
>>>
>>>> *****
>>>> To join, leave or search the confocal microscopy listserv, go to:
>>>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>>>> *****
>>>>
>>>> Is there any disadvantage to multiphoton for cultured cells  
>>>> (besides cost
>>>> and complexity)? Cell viability?  Phototoxicity?  Dave
>>>>
>>>> On Apr 23, 2011, at 11:21 AM, Kate Luby-Phelps wrote:
>>>>
>>>>
>>>>
>>>>> *****
>>>>> To join, leave or search the confocal microscopy listserv, go to:
>>>>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>>>>> *****
>>>>>
>>>>> Multiphoton has no clear advantage for cells in culture. For  
>>>>> cell culture
>>>>> samples,
>>>>> we use two photon  only to excite DAPI or other UV and near-UV  
>>>>> fluors
>>>>> since we
>>>>> had to make a choice between the Ti:S and a 405 laser on our  
>>>>> scope.
>>>>>
>>>>> Kate Luby-Phelps
>>>>>
>>>>>
>>>> Dr. David Knecht
>>>> Department of Molecular and Cell Biology
>>>> Co-head Flow Cytometry and Confocal Microscopy Facility
>>>> U-3125
>>>> 91 N. Eagleville Rd.
>>>> University of Connecticut
>>>> Storrs, CT 06269
>>>> 860-486-2200
>>>> 860-486-4331 (fax)
>>>>
>>>>
>>>>
>>>
>>>
>>> --
>>>
>>>
>>> George McNamara, PhD
>>> Analytical Imaging Core Facility
>>> University of Miami
>>>