Abstract: Diluents and methodologies for determining the concentration of total calcium in a clinical sample (e.g. whole blood, sera plasma urine cerebro spinal fluid) containing protein-bound calcium are disclosed. Methodologies include the steps of: (a) admixing the sample with a diluent, the diluent including effective amounts of a pH buffer and at least two agents capable of complexing free calcium in the sample, where the agent-calcium complex association constants ("log K" in base 10) each have a range from about 1.5 to about 7.0; and (b) contacting an aliquot of the diluted sample with a calcium-specific ion selective electrode. The response of the calcium-specific ion selective electrode is an indication of the concentration of total calcium in the sample.