14 May 2010 Influence of sample preparation and identification of subcelluar structures in quantitative holographic phase contrast microscopy
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Abstract
Digital holographic microscopy (DHM) provides label-free quantitative phase contrast with low demands on sample preparation. Nevertheless, for DHM measurements on fixed cells the mounting medium has to be considered while the phase contrast of living cells may be influenced by the used buffer solution. To quantify these effects, the maximum cell caused phase contrast and the visibility of the nucleoli were analyzed. A second aim of the study was to identify subcellular components in DHM phase contrast images. Therefore, comparative investigations using bright field imaging, DHM and fluorescence microscopy with 4',6- Diamidino-2-phenylindol (DAPI) staining were performed. DAPI-staining visualizes cell components containing DNA. The obtained results demonstrate exemplarily for two tumor cell lines that from DHM phase contrast images of fixed cells in phosphate buffer saline (PBS) cell thickness values are obtained which are comparable to living cells. Furthermore, it is shown that in many cases nucleus components can be identified only by DHM phase contrast.
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Björn Kemper, Björn Kemper, Lisa Schmidt, Lisa Schmidt, Sabine Przibilla, Sabine Przibilla, Christina Rommel, Christina Rommel, Angelika Vollmer, Angelika Vollmer, Steffi Ketelhut, Steffi Ketelhut, Jürgen Schnekenburger, Jürgen Schnekenburger, Gert von Bally, Gert von Bally, } "Influence of sample preparation and identification of subcelluar structures in quantitative holographic phase contrast microscopy", Proc. SPIE 7715, Biophotonics: Photonic Solutions for Better Health Care II, 771504 (14 May 2010); doi: 10.1117/12.854004; https://doi.org/10.1117/12.854004
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