Supplementary Figure 2: Comparative (non-TIRF/TIRF) excitation and emission patterns after micro-optics.
From: Single-shot super-resolution total internal reflection fluorescence microscopy
The response of each micro-optic as measured with fluorescence dye solution is shown for non-TIRF (entire objective back aperture illuminated, left column) and TIRF (annular illumination, middle column) excitation. Line scans corresponding to red lines in each image are shown in right column (blue: non-TIRF imaging; red: TIRF imaging). Although contrast using excitation microlenses is much better in TIRF than in non-TIRFl imaging, there is still residual background between excitation foci. This background is removed after adding pinholes; performance after pinholes (second row) and with emission microlenses (third row) is virtually identical whether exciting in TIRF or not. Note that these images were acquired with the galvanometric mirror held stationary, to visualize the emission around individual excitation foci. Scale bar: 2 μm. Experiments were repeated at least 3 times with similar results; representative data are shown.
