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Hi Kate

Do you really need/want two Tl:S lasers for this?  It may be better to  
use a visible and a Ti:S as I can foresee problems with continued  
photolysis of the cage with the probe beam if it's 2P... In our work  
measuring diffusion via 2P  we used an Ar-ion with a 2P photolysis  
spot that could be either stationary or moved with the probe using the  
same scanning system. For analysis the stationary spot (produced by  
our own custom optics coupling the Ti:S bean to the rear aperture  
independent of the scanner) was the best way to go. The trouble is not  
all microscope bodies can give you the room to a fit in a short pass  
dichroic to do this. In our  old Axiovert, we had to machine up a new   
coupling component to take a small dichroic on a slider. A  diagram of  
the layout is here:

Soeller, C. & Cannell, M.B. (1999) Two-photon microscopy: Imaging in  
scattering samples and three-dimensionally resolved flash photolysis.  
Microsc. Res. Tech. 47:182-195.

Cheers Mark


On 1/05/2011, at 1:06 AM, Kate Luby-Phelps wrote:

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> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> So far we have been using one laser, which means we miss the  
> beginning of the
> timecourse after photoactivation because it takes several seconds  
> for the Ti:S to
> tune and mode-lock.  So the experiments where 2P photoactivation has  
> been
> useful involve diffusion through gap junctions between cells in a  
> cluster or chain
> because the spread of the photoactivated dye between cells is  
> relatively slow.
> However, in June we will take delivery on a new 2P system with two  
> Ti:S lasers
> so we can do it simultaneously at two different wavelengths.
>
> Kate