Influence of sample preparation and identification of subcelluar structures in quantitative holographic phase contrast microscopy
2010
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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