Quantitative imaging of intact cardiac tissue using remote focusing microscopy

A. D. Corbett; R. A. B. Burton; G. Bub; T. Wilson

doi:10.1117/12.2076133

9 March 2015 Quantitative imaging of intact cardiac tissue using remote focusing microscopy

A. D. Corbett, R. A. B. Burton, G. Bub, T. Wilson

Proceedings Volume 9330, Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XXII; 93300R (2015) https://doi.org/10.1117/12.2076133
Event: SPIE BiOS, 2015, San Francisco, California, United States

Abstract

Remote focussing microscopy offers many advantages when acquiring volumetric data from living tissue. The all-optical means of refocussing does not agitate the specimen by moving either the stage or imaging objective. Aberrationcompensated imaging extends over volumes as large as 450 μm x 450 μm x 200 μm (X, Y and Z) allowing data to be collected from hundreds of cells. The speed with which refocussing can be achieved is limited only by the mechanical movement of a small (2 mm diameter) mirror. Using a pair of oblique imaging planes to rapidly acquire (<200ms) depth information temporally freezes residual tissue motion in the arrested heart.

This paper discusses the progress of remote focussing microscopy from a novel imaging technique to a reliable tool in the life sciences. Specifically, we describe recent efforts to achieve the accurate calibration of both distance and orientation within the imaging volume. Using a laser machined fluorescent specimen it is possible to identify, with high sensitivity, small (<1%) depth-dependent magnification changes which are a linear function of axial misalignment of the imaging objective. The sensitivity of the calibration procedure limits distortion to <1 μm over the entire imaging volume. This work finds direct application in identifying the microscopic effects of chronic disease in the living heart.

Citation Download Citation

A. D. Corbett, R. A. B. Burton, G. Bub, and T. Wilson "Quantitative imaging of intact cardiac tissue using remote focusing microscopy", Proc. SPIE 9330, Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XXII, 93300R (9 March 2015); https://doi.org/10.1117/12.2076133

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