Clearer view for TIRF and oblique illumination microscopy

In Total Internal Reflection Fluorescence (TIRF) microscopy, the sample is illuminated with an evanescent field that yields a thin optical section. However, its widefield detection has no rejection mechanism against out-of-focus blur from scattered light that can compromise TIRF images. Here I demonstrate that via structured illumination, out-of-focus blur can be effectively suppressed in TIRF microscopy, yielding strikingly clearer images. The same mechanism can also be applied to oblique illumination schemes that extend the reach of TIRF microscopy beyond the basal surface of the cell. The two imaging modes are used to image a biosensor, clathrin coated vesicles and the actin cytoskeleton in different cell types with improved contrast.