The osmotic potential of leaf sap was measured. A disc of Whatman GF/B glass micro-fibre paper was placed in the barrel of a 2 ml plastic syringe so that it covered the outlet hole. A fresh leaf was then put in the barrel, the plunger was re-inserted, and the tip of the syringe was sealed with Blu-Tack® (pressure-sensitive adhesive putty). The syringe was frozen in liquid nitrogen and, still sealed, was thawed to ambient temperature. When temperature equilibration was complete, the plunger and Blu-Tack were removed and the barrel of the syringe was placed in a 15 ml centrifuge tube, with its tip resting inside a 1.5 ml Ependorff tube. After centrifugation at 2500 g for 10 min at 4 °C, the osmolality of a 10 μl sample was measured by a calibrated vapour pressure osmometer