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David, I think from a theoretical viewpoint the multiphoton would have worse resolution because of the higher lambda (d = 900nm * 0.61 / NA), but better resolution due to smaller focal spot of 2P, so the resolution should be roughly equal to CLSM.

In practice (in my hands at least) CLSM images appear to have much better resolution. I would choose CLSM over 2P for the application you describe. 
Part of the problem as I see it comes from the wide excitation band of 2PE, meaning that you are exciting many fluorophores with a single wavelength, and you do not have this problem when using a "vis" laser.

Also, I think per unit area, in the focal spot, the 2PE will kill live cells faster than 1PE.

Cheers,   

Brian D Armstrong PhD
Assistant Research Professor
Light Microscopy Core 
Beckman Research Institute
City of Hope
Dept of Neuroscience
1450 E Duarte Rd
Duarte, CA 91010
626-256-4673 x62872

http://www.cityofhope.org/research/support/Light-Microscopy-Digital-Imaging/Pages/default.aspx


-----Original Message-----
From: Confocal Microscopy List [mailto:[log in to unmask]] On Behalf Of David Knecht charter
Sent: Friday, April 22, 2011 10:29 AM
To: [log in to unmask]
Subject: Live cell imaging with multiphoton confocal

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I have limited experience with multi-photon confocal microscopes.  I tried imaging live cells with one a while back and was unimpressed with the results as compared to a standard laser scanning instrument.  If you had a new single photon instrument and a new multi-photon instrument sitting side by side in your facility (and I presume some of you do), which would you choose for imaging  fixed cells or live cells in culture (not trying to go deep into tissue)?  Thanks- Dave

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)


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