Abstract
Matching specific cellular energy requirements with demand is a fundamental health determinant of individual cells, tissues, and organs. Two-photon excitation is currently an accepted method of quantifying cellular energetics however intensity based imaging alone cannot account for intracellular factors such as the lifetime differences in free and enzyme-bound NADH. As an alternative method to intensity based optical imaging, studies by Blinova et al. 2004', and Vishwasrao et al. 20052, suggest that time-resolved fluorescence lifetime imaging (FLIM) may provide a more accurate assesment of cellular metabolic function. We are currently in the process of comparing the capabilities of intensity and FLIM imaging as a means of assessing cellular metabolism. By using metabolic inhibitors and uncouplers to modify the metabolic state, we have observed six distinct lifetime populations of NADH in osteoblast-like 2T3 cells that represent free and enzyme bound forms of NADH. We are now characterizing multicell tumor spheroids, in vitro tumors which generate cell populations with varying metabolic rates in a spherically symetric system. By studying the various metabolic states located at different radii within the spheroid, we will compare NADH intensity imaging and FLIM in a realistic tumor environment.