We investigated the prognostic value of p16(INK4a) immunocytochemistry (ICC) analysis in 126 cases of newly diagnosed childhood acute lymphoblastic leukemia (ALL). The incidence of negative p16(INK4a) ICC was 38.1% and was more frequent in T-lineage ALL. Overall survival (OS) and event-free survival (EFS) were significantly higher in patients with positive p16(INK4a) ICC than in patients with negative ICC (6 years OS, 90% versus 63%, P =.0014; 6 years EFS, 77.8% versus 55%, P =.0033). The p16(INK4a) ICC remained a significant prognostic factor within the subgroup of B-precursor ALL. Multivariate analysis showed that negative p16(INK4a) ICC was an independent prognostic factor for OS (relative risk [RR], 3.38; P =.02) and EFS (RR, 2.49; P =.018). Sequential study showed that p16(INK4a) expression remained stable during first relapse in most patients. These findings indicate that p16(INK4a) ICC is an independent factor of outcome in childhood ALL.