An electromagnetic theory of imaging in fluorescence microscopy, and imaging in polarization fluorescence microscopy

Abstract

Imaging in fluorescence microscopy is analysed using a vectorial diffraction theory. Both conventional and confocal microscopy are considered. A fluorescent molecule is modelled as a radiating electric dipole. Images for particular orientations in fluorescence polarization microscopy are considered. We also average over all dipole orientations. In this case, two particular limiting cases are considered, corresponding to different depolarization relaxation times: the dipole can either freely rotate in space between excitation and emission, or is fixed in space. The image is different in each limiting case. If the dipole can freely rotate, the image after averaging is identical to that calculated assuming an isotropic point object.