Artifact-free, penetration-adjustable elliptical-mirror-based TIRF microscopy

Opt Express. 2018 Oct 1;26(20):26065-26079. doi: 10.1364/OE.26.026065.

Abstract

Evanescent field distribution in the focal region of the elliptical-mirror-based total-internal-reflection fluorescence (e-TIRF) microscopy is analyzed based on vectorial diffraction theory. The simulation demonstrates that the intensity of an evanescent field generated by elliptical mirror decreases exponentially with the penetration depth, and the polarization characteristic of the evanescent wave in various directions is given. We build up an e-TIRF microscope utilizing a focused hollow-cone illumination with all-direction and large range of incidence. The experiment shows the artifact effect can be well suppressed by using the azimuthal-direction illumination method. In addition, the penetration depth of the evanescent field can be controlled by adjusting the sizes of the aperture and obstruction with a large range.