Using Optical Spectroscopy to Measure Metabolic Activity in Islets
The proper functioning of pancreatic islet cells is essential for the production of insulin, which aids in regulating bodily glucose levels. Recent developments in islet transplantation have warranted the development of non-invasive methods to determine cell viability. This research project uses optical spectroscopy to measure metabolic activity in islets by: 1) evaluating NADH fluorescence and 2) measuring oxygen consumption through phosphorescence. Greater NADH fluorescence is a good indicator of viability because of its integral importance in the production of ATP, an important cellular free-energy donor. Since phosphorescence intensity is inversely proportional to oxygen concentration according to the Stern-Volmer law, phosphorescence can be used in oxygen sensing. Islets are placed in temperature-controlled glass wells with media and their NADH fluorescence is measured using a DMLP Leica microscope with epi-illumination. Baseline images of the fluorescence are taken using a CCD camera. These images are compared with images of the islets after treatment with low and high concentration glucose, which should initiate greater metabolic activity by stimulating insulin release. A final image is also taken after the application of sodium azide, which kills the cell and prevents further functional activity. To measure phosphorescence, a similar process is repeated on islets placed in wells painted with a thin film of phosphorescent dye such as platinum tetra (pentafluoro) phenyl porphyrin (PtTPFPP). The evaluation of the intensity of phosphorescence and NADH fluorescence in islets can provide vital information on the functional integrity of islets and their prospect for transplantation.
