Polarization Imaging of Cellular Autofluorescence

Intrinsic fluorophores and their associated biological processes exhibit dynamics on a wide range of timescales throughout the heterogeneous milieu of living cells. Conventional time-lapse autofluorescence intensity imaging is best suited for monitoring slow physiological functions such as changes in cellular morphology, cell migration, and intracellular distribution that take place on a timescale of seconds to minutes. In contrast, ultrafast (10^-12-10^-7 s) time-resolved fluorescence measurements can probe molecular dynamics, such as excited-state processes and rotational dynamics, which are acutely sensitive to the chemical structure, intermolecular interactions, and microenvironment of a given fluorophore. Time-resolved fluorescence and anisotropy measurements are uniquely suited for a detailed biophysical description of intrinsic fluorophores within the context of physiological and pathological changes in living cells or tissues. Consequently, multiparametric autofluorescence detection (i.e., intensity, color, lifetime, and polarization) provides the most complete description of intrinsically fluorescent biomolecules and their role in both the physiology and pathology of cells or tissues.