Fig. 3. Autonomous operation of the i-motif switch with time-programmed transient fluorescence as output. (a) Schematic depiction of the IFS-driven fluorescent i-motif switch. (b) Generation of self-regulated transient fluorescence under dynamic pH control by an IFS of 25 mM Tris (activator) and 300 mM β-BL (DD). The fluorescence signal (blue line) is shown together with the pristine pH curve (dashed gray line) of the pure IFS, and with the internal pH of the system in presence of the i-motif switch (black line). Measurements are carried out in water with 0.5 μM DNA, 100 mM NaCl and 0.1 mM EDTA. All fluorescence traces are collected at λ _em,max = 564 nm (λ _exc = 520 nm) and normalized to maximum intensity. Temporal programming of the fluorescence signal by (c) variation of the activator Tris at a constant DD concentration of 300 mM β-BL, and (d) variation of β-BL (DD) at a constant activator concentration of 25 mM Tris. Corresponding lifetime plots of the fluorescence signal in dependence of (e) Tris (300 mM β-BL) and (f) β-BL (25 mM Tris).