Characterizing Rotational Dynamics in Glassy Systems from Single Molecule Intensity Fluctuations

The dynamic heterogeneity exhibited by molecular rotations in glassy systems has long been characterized at the single molecule (SM) level by extracting timescales from linear dichroism (LD) collected via orthogonally polarized, wide-field fluorescence imaging. However, due to photons lost when collecting fluorescence images in this manner, localization precision is diminished relative to single channel collection. This makes examinations of dynamic heterogeneity as it manifests in translation measurements and the associated phenomenon of rotational-translational decoupling challenging to characterize as such measurements require high localization precision. Here, a method for extracting rotational dynamics of glassy systems at the SM level from intensity fluctuations of fluorescent probe molecules excited with circularly polarized light and collected in a single channel configuration is presented. Rotational dynamic timescales and degree of heterogeneity extracted from the LD and intensity approaches show good agreement, and the intensity-based approach is demonstrated to be robust across optical configurations, probe molecules, and in both polymeric and small molecule glass formers.