Mirrored pyramidal wells for simultaneous multiple vantage point microscopy

Authors

  • K.T. SEALE,

    1. Departments of Biological Sciences, Biomedical Engineering, Molecular Physiology & Biophysics and Physics, Vanderbilt Institute for Integrative Biosystems Research and Education, Vanderbilt University, VU Station B 351807, Nashville, TN 37235-1807, U.S.A.
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  • R.S. REISERER,

    1. Departments of Biological Sciences, Biomedical Engineering, Molecular Physiology & Biophysics and Physics, Vanderbilt Institute for Integrative Biosystems Research and Education, Vanderbilt University, VU Station B 351807, Nashville, TN 37235-1807, U.S.A.
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  • D.A. MARKOV,

    1. Departments of Biological Sciences, Biomedical Engineering, Molecular Physiology & Biophysics and Physics, Vanderbilt Institute for Integrative Biosystems Research and Education, Vanderbilt University, VU Station B 351807, Nashville, TN 37235-1807, U.S.A.
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  • I.A. GES,

    1. Departments of Biological Sciences, Biomedical Engineering, Molecular Physiology & Biophysics and Physics, Vanderbilt Institute for Integrative Biosystems Research and Education, Vanderbilt University, VU Station B 351807, Nashville, TN 37235-1807, U.S.A.
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  • C. WRIGHT,

    1. Departments of Biological Sciences, Biomedical Engineering, Molecular Physiology & Biophysics and Physics, Vanderbilt Institute for Integrative Biosystems Research and Education, Vanderbilt University, VU Station B 351807, Nashville, TN 37235-1807, U.S.A.
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  • C. JANETOPOULOS,

    1. Departments of Biological Sciences, Biomedical Engineering, Molecular Physiology & Biophysics and Physics, Vanderbilt Institute for Integrative Biosystems Research and Education, Vanderbilt University, VU Station B 351807, Nashville, TN 37235-1807, U.S.A.
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  • J.P. WIKSWO

    1. Departments of Biological Sciences, Biomedical Engineering, Molecular Physiology & Biophysics and Physics, Vanderbilt Institute for Integrative Biosystems Research and Education, Vanderbilt University, VU Station B 351807, Nashville, TN 37235-1807, U.S.A.
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John P. Wikswo. Tel: (615) 343-4124; fax: (615) 322-4977; e-mail: john.wikswo@vanderbilt.edu

Summary

We report a novel method for obtaining simultaneous images from multiple vantage points of a microscopic specimen using size-matched microscopic mirrors created from anisotropically etched silicon. The resulting pyramidal wells enable bright-field and fluorescent side-view images, and when combined with z-sectioning, provide additional information for 3D reconstructions of the specimen. We have demonstrated the 3D localization and tracking over time of the centrosome of a live Dictyostelium discoideum. The simultaneous acquisition of images from multiple perspectives also provides a five-fold increase in the theoretical collection efficiency of emitted photons, a property which may be useful for low-light imaging modalities such as bioluminescence, or low abundance surface-marker labelling.

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