Figure 2.

Optimization of tFCS spatial sensitivity with side-illumination, and expected resolution in presence of tracking localization error. (A and B) A schematic of the centered- (A) and side-illumination (B) configurations is shown. Top: the relative positions of the tracking (green) and probe (red) beams are shown. Bottom: the X axis cross-section of the probe laser intensity profile seen by the probe dye is shown. The green cross represents the lock point of the feedback (at the center of tracking laser circular orbit) with respect to which the reference dye is immobile. The green and red stars represent the average position of the reference and probe dyes in the beam, respectively. (C) The illustration shows the effect of finite-bandwidth tracking, in which the reference dye is imperfectly maintained at the lock point, which results in a residual jitter of the probe dye in the probe beam frame. (D and E) Simulated tFCS signals (mean $\pm$ 95% confidence interval) of molecules with a reference-to-probe dye root mean-square distance (RMSD) of 0, 17, 35, or 87 nm are shown, using either the centered- (D) or the side-illumination (E) configuration. In the side-illumination geometry, the reference-to-probe beam offset was set to $1.0w$, where $w=310\text{nm}$. Asterisk color indicates the shortest resolvable reference-to-probe RMSD. (F) The same simulation as that in (E) is shown, but this time taking into account imperfect feedback (100 nm RMS localization error). To see this figure in color, go online.