Re: Changing system from NIR (two-photon) to UV

Posted by Zoltan on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Changing-system-from-NIR-two-photon-to-UV-tp1694402p1694838.html

Hi Esteban,

Just a few thoughts, because I've never attempted such a swap of lasers: for DAPI, we use a 405 nm laser, and we found no need for real UV.  However, even with this near-UV line, the light path is separate from the visible light paths and the near-UV optics contains correction pinhole lenses (one pinhole lens for each objective lens) in order to be able to align the visible and near-UV generated confocal images (even a slight misalignment of these corrective elements results in a 1 to 5 micron misalignment between the two images).  We also image nuclei regularly and the light paths have to be well aligned if we want to have images of any use, thus I very much doubt that you would get away with a simple swap.
  I hope this helps a bit, I'm sure our more experienced list members will fill us in with great info soon. Best regards,

Zoltan 

On Tue, Dec 23, 2008 at 4:47 PM, G. Esteban Fernandez <[hidden email]> wrote:

Hello all,

 

On our Zeiss LSM 510 META two-photon system we have a Coherent Chameleon NIR laser that may need major repair.  Coherent has been very good in providing remote technical assistance to us but we're at a point where a service visit is required and we're not under contract.  Since we do more shallow imaging of UV dyes like DAPI on that system that two-photon imaging, we would replace the NIR laser with UV if it is more cost-effective than repairing the NIR.  Zeiss says a switch from NIR to UV would require a major (and pricey) overhaul of the optics.  I don't know what that entails but it is understandable given the huge difference in wavelength.  However, I wonder how badly out of focus or aberrated the UV image would actually be with the optics that are already in place if we put a UV laser in there in place of the NIR; we do have a UV laser dichroic.  Might it be good enough for at least localizing nuclei? (I do know how to do non-confocal UV imaging on the system)  Can people advise on an NIR-to-UV swap of lasers?

 

Thanks,

Esteban

--
G. Esteban Fernandez, Ph.D.
Associate Director
Molecular Cytology Core Facility
University of Missouri
120 Bond Life Sciences Center
Columbia, MO  65211

http://www.biotech.missouri.edu/mcc/

573-882-4895
573-884-9395 fax

--

Zoltan Cseresnyes
Facility manager, Imaging Suite