The Calcium Calibration Buffer Kits are designed to help determine the dissociation constant (K_d) of fluorescent Ca²⁺ indicators using a fluorometer or quantitative imaging system. The Kd of an indicator can be calculated from a plot generated by scanning the excitation or emission of the indicator in the presence of eleven different Ca²⁺ concentrations. Calcium Calibration Kit #1 contains 50 mL of 10 mM K₂EGTA and 50 mL of 10 mM CaEGTA. Both solutions contain 100 mM KCl and 30 mM MOPS, pH 7.2 and are prepared in deionized water. These stock solutions can be blended to make buffers having free Ca2+ ranging from 0 µM to 39 µM.