Fig. 3. Fluorescence pH titration of CF and CR co-assembled on rec-Ori (rec-CF5/CR5) (1.6 nM). (a) The structure of rec-CF5/CR5. (b and c) Fluorescence emission spectral changes of (b) CF (excited at 470 nm) and (c) CR (excited at 530 nm) at various pH in the range from 3 to 10. (d) A plot for the ratio of emission intensity at 518 nm excited at 470 nm, corresponding to CF, to that at 582 nm excited at 530 nm, corresponding to CR, for rec-CF5/CR5 at various pH in the range from 3 to 10. The emission intensity ratios (518/582) were calculated by dividing the fluorescence intensity at 518 nm with excitation at 470 nm by the fluorescence intensity at 582 nm with excitation at 530 nm. The pK_a value was estimated to be 7.1 ± 0.1 at ambient temperature. The details of the buffer used for each pH condition were shown in Experimental procedures.