Although alterations in the shape and size of subcellular organelles are important markers of cell death, these important morphological changes cannot be directly quantified in living cells by conventional light microscopy. Quantitative light scattering provides an appealing solution to this problem. This technique is noninvasive, and is sensitive to the dimensions of particles with size on the order of the wavelength. Existing light scattering methods, which have been successfully utilized in flow cytometry, are spectroscopic in nature and do not permit sample imaging. Preserving spatial information is crucial to correlating the source of the scatter changes with a given cell or organelle. To this end, the investigators propose to combine light scattering spectroscopy with imaging. This novel technology, OSM, will produce the first images in which alterations in organelle morphology can be tracked in living cells in real time. The preliminary data attest to the feasibility and sensitivity of OSM. Notably, the investigators were able to detect an intracellular optical scatter change early during apoptosis, long before it was discernable by other light microscopic methods, such as DIC. However, the biological events responsible for this scatter change remain to be identified. Elucidating the relationship between the optical scatter changes and cellular activity is the principal challenge in developing OSM. If this challenge is met, OSM has the potential to transform significantly the approach to non-invasive monitoring of intracellular activity, and holds promise to fundamentally enhance high resolution light microscopy. In this proposal, the investigators present a rational strategy and step-by- step approach to implement and validate OSM. In this initial phase of development, the investigators will use OSM to study mitochondrial dysfunction during cell death. The long-term goal is to develop OSM as a tool for functional microscopy.
| Zheng, Jing-Yi; Tsai, Yien-Che; Kadimcherla, Pradeep et al. (2008) The C-terminal transmembrane domain of Bcl-xL mediates changes in mitochondrial morphology. Biophys J 94:286-97 |
| Boustany, Nada N; Tsai, Yien-Che; Pfister, Bryan et al. (2004) BCL-xL-dependent light scattering by apoptotic cells. Biophys J 87:4163-71 |
| Boustany, Nada N; Drezek, Rebekah; Thakor, Nitish V (2002) Calcium-induced alterations in mitochondrial morphology quantified in situ with optical scatter imaging. Biophys J 83:1691-700 |
