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Hi David,
those powers could potentially be made to work at a pinch, if you were prepared to limit yourself to a very small field of view, but in general around 200-500mW, or possibly even higher is where you want to be. The 405 probably already has enough power as is. Are you planning on doing PALM (fluorescent proteins) or STORM(organic dyes)? In my opinion, the first laser to replace would be the 640, as this is where you will get the most mileage. A good 200mW laser at 640nm should go for somewhere in the vicinity of 7-8K. If you are prepared to go longer, reasonable but decidedly no-frills 671nm lasers at ~ 1W can be had from Viasho for about 2K (although there are some caveats with using this wavelength). A good 561/568nm laser would probably be the next step - although these are typically more expensive (12-15k). You will also want to replace the TIRF coupler (probably with something home built) as the stock one spreads the laser light out over too large a field of view to be useful (ideally you want to just illuminate an area the size of your camera FOV, potentially even smaller). Depending on how your current focusing is implemented, you might want to consider a piezo-focusser and/or some form of 'perfect focus' scheme.
I've got a custom system based on a Nikon Ti running down here in New Haven - if you want to come down for a look, drop me an email at [log in to unmask]
cheers,David
On Friday, 16 January 2015 4:20 PM, "Knecht, David" <[log in to unmask]> wrote:
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We are looking into setting up a super-resolution system using our existing laser combiner and Nikon TI TIRF arm setup to bring the light to the sample. The system is run with Micro-Manager, so QuickPalm seems like a good place to start. We currently have 50W 405/75 mW 488/ 50mW 561/ 40mW 640 diode lasers on the system. I have no experience with super resolution at all yet but I have been told that this is not enough laser power for QuickPalm or STORM. Some funds from the university have suddenly appeared and I need to respond rapidly so I would like to know if people have suggestions as to specifically what lasers I should be looking at to add to the system and what they are likely to cost. Any hints as to other problems I am not expecting that will need $$$ to solve would be helpful as well. Thanks- Dave
Dr. David Knecht
Professor of Molecular and Cell Biology
Core Microscopy Facility Director
University of Connecticut
Storrs, CT 06269
860-486-2200
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