AbstractRecently, the field of complexometric titration significantly progressed with the advent of ionophore‐based titration reagents. We present here for the first time an ionophore‐based heterogeneous optical titration method for calcium. The all‐in‐one reagent was composed of chromoionophore, ion‐exchanger and calcium ionophore in dichloromethane (DCM). The endpoint of this reverse titration scheme was determined by the color change of the DCM phase (blue to red). The ion‐exchange theory borrowed from ISE membranes was used to simulate the titration curve in a simplified manner. The phase separation speed during titration was found much faster at and beyond the endpoint. A mechanism was proposed for this phenomenon, with zeta‐potential measurements on ion‐selective nanospheres to support the hypothesis. Calcium titration in a urine sample was also successfully demonstrated and compared with EDTA titration with potentiometric ion‐selective electrodes as endpoint indicator. The proposed reagent serves as an alternative to the classical EDTA.