Figure 4.

Ion measurement using a pH-correctable paradigm. (a) Design and working principle of the pH-correctable Ca²⁺ reporter, CalipHluor. (b) Structure of the Ca²⁺ sensing fluorophore, Rhod-5F. 3D calibration surfaces of (c) D/A and (d) O/R as a function of pH and Ca²⁺. (e) Image analysis workflow for heatmaps of free Ca²⁺ in lysosomes. The pH map is obtained from the D/A versus pH curve as in 1. Then, the K_d map is generated from a function describing pH versus K_d as is 2. The Ca²⁺ map is obtained from the O/R map and K_d map using eq 3.