Objective
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ============================================================ Can anyone suggest an high numerical aperature 20 or 25x infinity objective lens that works in the UV range to be be used for Ca++ ratioing using Fura2 dye? |
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Hello Christie, What microscope stand do you have? It really does matter, due to final corrections. If you have a Zeiss stand, then these objectives have good transmission in the UV and very high N.A., but it is not very well corrected for Chromatic or Spherical aberrations: Zeiss Fluar 20x/0.75 https://www.micro-shop.zeiss.com/de/de_de/objektive.php?cp_sid=&f=od&p[]=440145-0000-000 - (this one seems to have better UV transmission) Found using: If you have an Olympus stand this objective is better corrected for Chromatic or Spherical aberrations, but doesn't have the resolving power of the Zeiss objective due to lower N.A.: But this one ROCKS! Having the best correction of all of the other objectives listed: Found using: If you have a Nikon stand: S Fluar 20x/0.75 is most likely your best bet... Found using: If you have a Leica stand: Maybe a HCX PL FLUOTAR 20x/0.50 POL will work for you. Found using: http://www.leica-microsystems.com/products/light-microscopes/accessories/objectives/Good luck finding what you need! Pete On Sep 15, 2010, at 17:55 PM, Christie DeVille wrote:
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In reply to this post by Christie DeVille
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ============================================================ Dear Christie, we had worked quite a lot on that issue, though this was back in the 90s. The question is, whether you would like to use the Argon laser UV wavelengths optimal for Fura-2, i.e. 334nm and 364nm, or whether you are prepared to go for something less optimal, e.g. 351nm and 364nm. In the latter case, quite a number of objectives will do the job, though not at confocal precision unless used with adaptive optics. In the former case, it's really a problem and the only objectives that possibly might be suitable are the Zeiss Ultrafluars. However, on a Zeiss microscope, you would also need a special tube lens. I refer to our following publications: P.J. Helm, O. Franksson, and K. Carlsson (1995), A confocal scanning laser microscope for quantitative ratiometric 3D measurements of [Ca2+] and Ca2+ diffusions in living cells stained with Fura-2, Pflügers Archiv - European Journal of Physiology, Vol. 429, pp. 672-681 and PJ Helm, A Patwardhan, and EMM Manders (1997), A study of the precision of confocal, ratiometric, Fura-2 based measurements, Cell Calcium 22(4):287-298 Microscope optically, you might have an easier life with Indo-1, which demands one UV wavelength, only. The situation is different with a multi photon approach, where it is really a problem to separately excite the two Fura-2 modifications. Best wishes Johannes ---------------------------- Original Message ---------------------------- Subject: Objective From: "Christie DeVille" <[hidden email]> Date: Wed, September 15, 2010 17:55 To: [hidden email] -------------------------------------------------------------------------- ============================================================ To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ============================================================ Can anyone suggest an high numerical aperature 20 or 25x infinity objective lens that works in the UV range to be be used for Ca++ ratioing using Fura2 dye? > ============================================================ > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ============================================================ > > Can anyone suggest an high numerical aperature 20 or 25x infinity > objective > lens that works in the UV range to be be used for Ca++ ratioing using > Fura2 dye? > -- P. Johannes Helm, M.Sc. PhD Seniorengineer CMBN University of Oslo Institute of Basic Medical Science Department of Anatomy Postboks 1105 - Blindern NO-0317 Oslo Voice: +47 228 51159 Fax: +47 228 51499 WWW: folk.uio.no/jhelm |
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you should look also at the following two publications from us in the late 90s with confocal ratiometric fura-2 or 2-P fura-2 measurements: A modified confocal laser scanning microscope allows fast ultraviolet ratio imaging of intracellular Ca2+ activity using Fura-2. Nitschke R, Wilhelm S, Borlinghaus R, Leipziger J, Bindels R, Greger R. Pflugers Arch.
1997 Mar;433(5):653-63. and Ricken S, Leipziger J, Greger R, Nitschke R. J Biol Chem. 1998 Dec 25;273(52):34961-9.regards Roland ___________________________ Nitschke, Roland Dr. Life Imaging Center Centre of Systems Biology Albert-Ludwigs-University Freiburg Habsburgerstr.49 D-79104 Freiburg Germany ___________________________ E-mail: [hidden email] phone: 49-761-2032934 or 2902 fax: 49-761-2032941 http://www.imaging.uni-freiburg.de/ Am 16.09.2010 11:41, schrieb P. Johannes Helm: ============================================================ To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ============================================================ Dear Christie, we had worked quite a lot on that issue, though this was back in the 90s. The question is, whether you would like to use the Argon laser UV wavelengths optimal for Fura-2, i.e. 334nm and 364nm, or whether you are prepared to go for something less optimal, e.g. 351nm and 364nm. In the latter case, quite a number of objectives will do the job, though not at confocal precision unless used with adaptive optics. In the former case, it's really a problem and the only objectives that possibly might be suitable are the Zeiss Ultrafluars. However, on a Zeiss microscope, you would also need a special tube lens. I refer to our following publications: P.J. Helm, O. Franksson, and K. Carlsson (1995), A confocal scanning laser microscope for quantitative ratiometric 3D measurements of [Ca2+] and Ca2+ diffusions in living cells stained with Fura-2, Pflügers Archiv - European Journal of Physiology, Vol. 429, pp. 672-681 and PJ Helm, A Patwardhan, and EMM Manders (1997), A study of the precision of confocal, ratiometric, Fura-2 based measurements, Cell Calcium 22(4):287-298 Microscope optically, you might have an easier life with Indo-1, which demands one UV wavelength, only. The situation is different with a multi photon approach, where it is really a problem to separately excite the two Fura-2 modifications. Best wishes Johannes ---------------------------- Original Message ---------------------------- Subject: Objective From: "Christie DeVille" [hidden email] Date: Wed, September 15, 2010 17:55 To: [hidden email] -------------------------------------------------------------------------- ============================================================ To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ============================================================ Can anyone suggest an high numerical aperature 20 or 25x infinity objective lens that works in the UV range to be be used for Ca++ ratioing using Fura2 dye?============================================================ To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ============================================================ Can anyone suggest an high numerical aperature 20 or 25x infinity objective lens that works in the UV range to be be used for Ca++ ratioing using Fura2 dye? |
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