Here we report a simple screening system using hematoxylin staining (HS) of the root apex. It allowed rapid classification into different aluminum (Al) tolerance from 65 cultivars within one week. Using this system, we selected the most Al-tolerant (Jayae-2) and -sensitive (Pum-2) barley (Hordeum vulgare L.) The results show that the different responses in Al-induced growth inhibition, Al accumulation and expression of plasma membrane (PM) H+-ATPase in root apices of selected two cultivars. It showed strongly Al-induced growth inhibition in a dosedependant manner only in Pum-2 but not in Jayae-2. Aluminum accumulation in root apices (10 mm) was significantly higher in Pum-2 only. The H+-ATPase expression of PM vesicles by western blotting was decreased in Pum-2 but not in Jayae-2 treated with 20 μM Al for 24 h. These finding indicate to screen from our system is rapid and reliable and to sustain the expression of PM H+-ATPase at translational level is an important role in root growth as affected by Al.