Abstract
The origins of side scattering from a fibroblast and cervical cell line were determined by comparing side-scatter images with images stained for lysosomes, nuclei, and mitochondria on a cell by cell basis. Lysosomes or nuclei are the most efficient type of scatterer depending on the cell type and incident light polarization. The relative scattering efficiencies of lysosomes and mitochondria were the same for both cell lines, while the scattering efficiencies of the nuclei differed. The percent of 90° scattering from the nucleus, mitochondria, and lysosomes as well as the group of other internal cellular objects was estimated. The nucleus was the largest contributor to side scatter in the cervical carcinoma cells. The contributions of lysosomes, mitochondria, the nucleus, and particles unstained by either Hoechst, LysoSensor or MitoTracker ranges from ∼20% to ∼30% in fibroblast cells. The contribution of lysosomes to side scatter was much stronger when the incident light was polarized perpendicular to the scattering plane than when the polarization of the side scatter laser was parallel to the scattering plane. This dependence on side scatter polarization indicates that lysosomes contain scattering structures that are much smaller than the wavelength of light used in the measurements (785 nm). In conclusion, mitochondria were not found to be either the most efficient scatterer or to have the largest contribution to scattering in either cell line, in contrast to previous reports. Rather lysosomes, nuclei and unknown particles all have significant contributions to 90° scattering and the contributions of some of these particles can be modulated by changing the polarization of the incident light.
Highlights
Light scattering measurements provide information about cells without the need for stains or other perturbation that might affect the cell or hinder subsequent measurements
Higher resolution images of mitochondria in fibroblast cells show a thread-like pattern with some dense areas within the cytoplasm [18] and our results are consistent with this result
Lysosomes are more efficient side scatterers than mitochondria, especially when the incident light is polarized perpendicular to the scattering plane
Summary
Light scattering measurements provide information about cells without the need for stains or other perturbation that might affect the cell or hinder subsequent measurements. Work in the 1970’s demonstrated that light scattering properties are related to cell morphology [1, 2]. This fact has motivated the development of in vivo light scatter techniques for nonintrusive diagnosis of precancerous conditions. Work by Beauvoit et al demonstrated that mitochondria are a major contributor to light scattering from liver cells [9]. These results can not be extrapolated to other cell types. Mitochondria are much less of the cell volume. In human skin fibroblasts, mitochondria are 2.4% and lysosomes are more than 2.7% of the cell volume [11]
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