EGTA purity and the buffering of calcium ions in physiological solutions

Results are presented for the purity of three commercial brands of the Ca chelator EGTA [ethyleneglycol-bis(beta-aminoethylether)-N,N'-tetraacetic acid] as determined with pH-metric and oxalate precipitation methods and other techniques. The effect that the observed degree of hydration (2-4%) has on calculated values for [Ca2+] in Ca-EGTA solutions is described with reference to a theoretical dose-response relationship. In addition to the corrections for purity, the adjustments necessary to allow for ionic strength and the correction of pH readings to yield true [H+] activity or concentration values are described. These several corrections result in both a shift and a shape change for the dose-response curve compared with the curve calculated without these corrective procedures. Some authors have already taken some of these aspects into account, but this has not generally been made clear in the literature heretofore. The points raised may help to account for the wide variation of reported [Ca2+] ranges for Ca-sensitive cellular processes.