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

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 i...

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Bibliographic Details
Published in:Journal of Biomedical Optics 2008-01, Vol.13 (1), p.014011-0140110
Main Authors: Shribak, Michael, LaFountain, James, Biggs, David, Inoue, Shinya
Format: Article
Language:English
Subjects:
Algorithms
Anisotropy
Architecture
birefringence
Cranes
differential interference contrast (DIC) microscopy
dry mass
Equipment Design
Equipment Failure Analysis
Image contrast
Image Enhancement - instrumentation
Image Enhancement - methods
Image Interpretation, Computer-Assisted - instrumentation
Image Interpretation, Computer-Assisted - methods
Interference
Microscopy, Polarization - instrumentation
Microscopy, Polarization - methods
optical path gradient
phase microscopy
Polarization
polarized microscopy
Reproducibility of Results
Sensitivity and Specificity
Staining
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Summary: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.
ISSN:1083-3668
1560-2281
DOI:10.1117/1.2837406