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Methanol immersion reduces spherical aberration of water dipping lenses at long wavelengths used in multi-photon laser scanning microscopy

Norris, Greg and Gebril, Ayman M and Ferro, Valerie and McConnell, Gail (2012) Methanol immersion reduces spherical aberration of water dipping lenses at long wavelengths used in multi-photon laser scanning microscopy. Biomedical Optics Express, 3 (12). pp. 3314-3324. ISSN 2156-7085

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    Abstract

    Dipping objectives were tested for multi-photon laser scanning microscopy, since their large working distances are advantageous for thick specimens and the absence of a coverslip facilitates examination of living material. Images of fluorescent bead specimens, particularly at wavelengths greater than 850 nm showed defects consistent with spherical aberration. Substituting methanol for water as the immersion medium surrounding the beads corrected these defects and produced an increase in fluorescence signal intensity. The same immersion method was applied to two representative biological samples of fixed tissue: mouse brain labeled with FITC for tubulin and mouse gut in which the Peyer’s patches were labeled with Texas Red bilosomes. Tissue morphology was well preserved by methanol immersion of both tissues; the two-photon-excited fluorescence signal was six times higher than in water and the depth of penetration of useful imaging was doubled. No modification of the microscope was needed except the provision of a ring to retain a sufficient depth of methanol for imaging.

    Item type: Article
    ID code: 42199
    Keywords: multi-photon laser scanning microscopy, wavelengths , fluorescent beads, tissue morphology, methanol, Pharmacy and materia medica, Biotechnology, Atomic and Molecular Physics, and Optics
    Subjects: Medicine > Pharmacy and materia medica
    Department: Faculty of Science > Strathclyde Institute of Pharmacy and Biomedical Sciences
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    Depositing user: Pure Administrator
    Date Deposited: 24 Nov 2012 12:36
    Last modified: 06 Sep 2014 13:59
    URI: http://strathprints.strath.ac.uk/id/eprint/42199

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