Effect of rotational diffusion in an orientational potential well on the point spread function of electric dipole emitters.

Abstract

A study is presented of the point spread function (PSF) of electric dipole emitters that go through a series of absorption-emission cycles while the dipole orientation is changing due to rotational diffusion within the constraint of an orientational potential well. An analytical expression for the PSF is derived valid for arbitrary orientational potential wells in the limit of image acquisition times much larger than the rotational relaxation time. This framework is used to study the effects of the direction of incidence, polarization, and degree of coherence of the illumination. In the limit of fast rotational diffusion on the scale of the fluorescence lifetime the illumination influences only the PSF height, not its shape. In the limit of slow rotational diffusion on the scale of the fluorescence lifetime there is a significant effect on the PSF shape as well, provided the illumination is (partially) coherent. For oblique incidence, illumination asymmetries can arise in the PSF that give rise to position offsets in localization based on Gaussian spot fitting. These asymmetries persist in the limit of free diffusion in a zero orientational potential well.