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Research Papers

Orientation-independent differential interference contrast microscopy and its combination with an orientation-independent polarization system

[+] Author Affiliations
Michael Shribak

Marine Biological Laboratory, 7 MBL Street, Woods Hole, Massachusetts 02543

James LaFountain

University at Buffalo, Department of Biological Sciences, 657 Cooke Hall, Buffalo, New York 14260

David Biggs

AutoQuant Imaging, Inc., 105 Jordan Road, Troy, New York 12180

Shinya Inouè

Marine Biological Laboratory, 7 MBL Street, Woods Hole, Massachusetts 02543

J. Biomed. Opt. 13(1), 014011 (February 12, 2008). doi:10.1117/1.2837406
History: Received April 28, 2007; Revised October 01, 2007; Accepted October 23, 2007; Published February 12, 2008
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We describe a combined orientation-independent differential interference contrast OI-DIC and polarization microscope and its biological applications. Several conventional DIC images were recorded with the specimen oriented in different directions followed by digital alignment and processing of the images. Then the obtained images are used for computation of the phase gradient magnitude and azimuth distribution and, further, the phase image. The OI-DIC images were obtained using optics having numerical aperture (NA) 1.4, thus achieving a level of resolution not previously achieved with phase contrast or interference microscope. The combined system yields two complementary phase images of thin optical sections of the specimen: distribution of refractive index and distribution of birefringence due to anisotropy of the cell structure. For instance, in a live dividing cell, the OI-DIC image clearly shows the detailed shape of the chromosomes, while the polarization image quantitatively depicts the distribution of birefringent microtubules in the spindle, both without any need for staining or other modifications of the cell. We present pseudo-color combined images of a crane fly spermatocyte at diakinesis and metaphase of meiosis I. Those images provide clear evidence that the proposed technique can reveal fine architecture and molecular organization in live cells without perturbation associated with staining or fluorescent labeling.

Figures in this Article
© 2008 Society of Photo-Optical Instrumentation Engineers

Citation

Michael Shribak ; James LaFountain ; David Biggs and Shinya Inouè
"Orientation-independent differential interference contrast microscopy and its combination with an orientation-independent polarization system", J. Biomed. Opt. 13(1), 014011 (February 12, 2008). ; http://dx.doi.org/10.1117/1.2837406


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