After the discovery of ion-mediated changes in xylem hydraulic resistance a few years ago, a number of research papers were published that related ion-mediated flow changes in the xylem to various aspects of whole plant functioning and evolutionary diversification of vascular cells. Ion-mediated changes in xylem hydraulic resistance are commonly quantified as the percentile change in hydraulic resistance, relative to the hydraulic resistance measured using a reference fluid, usually (ultra) pure deionized water. In this research the impact was investigated of the complete absence of all ions in deionized water compared with reference fluids containing a minimal amount of free calcium on the quantification of ion-mediated flow changes in stem segments of Chrysanthemum (Dendranthema×grandiflorum Tzvelev) and Prunus L. (Prunus laurocerasus L.). The addition of 10 mM KCl to deionized water significantly increased flow rate in Chrysanthemum (17–24%) and Prunus L. (16%). The addition of 1 mM CaCl2 to the reference fluid reduced this KCl-mediated increase in flow rate to 1–2% in both species. 1 mM Ca2+ is within the lower range of Ca2+-concentrations normally measured in xylem sap of many plant species, and three times lower than the original Ca2+-concentration measured in the xylem sap of Chrysanthemum plants used for the present measurements. The present results indicate that the complete removal of cations from the xylem fluid with deionized water causes the major part of the ion-mediated flow change previously reported in the xylem of plants. It is concluded that the use of deionized water as a reference fluid should be avoided. Earlier proposed relationships between ion-mediated changes and water flow in xylem of plants should be re-evaluated if they were based on deionized water as the reference fluid.