Eliminating unwanted far-field excitation in objective-type TIRF. Part II. combined evanescent-wave excitation and supercritical-angle fluorescence detection improves optical sectioning.

ABSTRACT: Azimuthal beam scanning makes evanescent-wave (EW) excitation isotropic, thereby producing total internal reflection fluorescence (TIRF) images that are evenly lit. However, beam spinning does not fundamentally address the problem of propagating excitation light that is contaminating objective-type TIRF. Far-field excitation depends more on the specific objective than on cell scattering. As a consequence, the excitation impurities in objective-type TIRF are only weakly affected by changes of azimuthal or polar beam angle. These are the main results of the first part of this study (Eliminating unwanted far-field excitation in objective-type TIRF. Pt.1. Identifying sources of nonevanescent excitation light). This second part focuses on exactly where up beam in the illumination system stray l

SUBMITTER: Brunstein M 

PROVIDER: S-EPMC4026779 | biostudies-literature | 2014 Mar

REPOSITORIES: biostudies-literature