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Digital Holographic Microscopy: Quantitative Phase Imaging and Applications in Live Cell Analysis

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Handbook of Coherent-Domain Optical Methods

Abstract

The analysis of complex processes in living cells creates a high demand for fast and label-free methods for online monitoring. Widely used fluorescence methods require specific labeling and are often restricted to chemically fixated samples. Thus, methods that offer label-free and minimally invasive detection of live cell processes and cell state alterations are of particular interest. In combination with light microscopy, digital holography provides label-free, multi-focus quantitative phase imaging of living cells. In overview, several methods for digital holographic microscopy (DHM) are presented. First, different experimental setups for the recording of digital holograms and the modular integration of DHM into common microscopes are described. Then the numerical processing of digitally captured holograms is explained. This includes the description of spatial and temporal phase shifting techniques, spatial filtering based reconstruction, holographic autofocusing, and the evaluation of self-interference holograms. Furthermore, the usage of partial coherent light and multi-wavelength approaches is discussed. Finally, potentials of digital holographic microscopy for quantitative cell imaging are illustrated by results from selected applications. It is shown that DHM can be used for automated tracking of migrating cells and cell thickness monitoring as well as for refractive index determination of cells and particles. Moreover, the use of DHM for label-free analysis in fluidics and micro-injection monitoring is demonstrated. The results show that DHM is a highly relevant method that allows novel insights in dynamic cell biology, with applications in cancer research and for drugs and toxicity testing.

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Acknowledgments

Financial support by the German Ministry for Education and Research (BMBF) is gratefully acknowledged.

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Authors and Affiliations

  1. Center for Biomedical Optics and Photonics Muenster, University of Münster, Münster, Germany

    Björn Kemper, Patrik Langehanenberg, Sebastian Kosmeier, Frank Schlichthaber, Christian Remmersmann & Gert von Bally

  2. Biomedical Technology Center, University of Münster, Münster, Germany

    Christina Rommel, Christian Dierker & Jürgen Schnekenburger

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  1. Björn Kemper
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  2. Patrik Langehanenberg
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  3. Sebastian Kosmeier
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  4. Frank Schlichthaber
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  6. Gert von Bally
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  7. Christina Rommel
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  8. Christian Dierker
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  9. Jürgen Schnekenburger
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Correspondence to Björn Kemper .

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  1. , Chair of Optics and Biophotonics, Saratov State University, Astrakhanskaya str. 83, Saratov, 410012, Russia

    Valery V. Tuchin

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Kemper, B. et al. (2013). Digital Holographic Microscopy: Quantitative Phase Imaging and Applications in Live Cell Analysis. In: Tuchin, V. (eds) Handbook of Coherent-Domain Optical Methods. Springer, New York, NY. https://doi.org/10.1007/978-1-4614-5176-1_6

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