ABSTRACT
This chapter demonstrates how to check polarizers, how to measure polarized absorption of chromophores embedded into an anisotropic media, and how to measure emission/excitation polarization spectra. Measuring emission polarization is more routine and a typical spectrofluorometer should be equipped with polarizers. Excitation with linearly polarized light will preferentially select molecules that have an absorption transition moment parallel to the direction of light polarization. The polarization of emission light can be measured and may provide very important information about molecular orientation and changes in molecular orientation between the act of absorption and act of emission. It is important to stress that the observed fluorescence signal at the magic angle is independent of sample polarization and will not be affected by depolarization processes like rotational diffusion. Measurements of fluorescence anisotropy require the excitation with polarized light and the detection of fluorescence intensity observed through a parallel and perpendicular oriented polarizer as compared to the excitation light polarization.
