2.7 Microelectrode ion flux estimation (MIFE)
Al-induced net fluxes of HPO42- were measured from elongation zone of three-day-old barley seedlings using MIFE (Chen et al., 2007). LIX for HPO42- was prepared using N,N-Bis(salicylidene)-o-phenylenediamine vanadium (IV) oxide complex (Sigma, Switzerland) as ionophore according to Ganjali, Mizani, & Salavati-Niasari (2003). The seedling was transferred to a 5-ml Perspex measuring chamber with 2.5 ml MIFE basal solution (500 μM KCl, 100 μM CaCl2, pH 4.3), one hour prior to flux measurement. Net ion flux measurement commenced in the basal solution for 10 min and followed by 30 min in 25 μM Al3+ treatment by adding 2.5 ml treatment solution (50 μM AlCl3, 500 μM KCl, 100 μM CaCl2, pH 4.3). The average flux in control (0-10 min), transient (11 to 15 min) and steady-state (30-40 min) was calculated.