Phosphorylation detected in HEK293 cells after treatment with ionising radiation (IR) (10Gy). The antibody was incubated with the non-phosphorylated peptide (40 μg/ml) for 1 hour before addition to the nitrocellulose membrane. The antibody was used at 1 μg/ml in 5 % skimmed milk / TBS-Tween and incubated with the membrane for 1 hour at room temperature. Binding of the primary antibody was detected using rabbit peroxidase conjugated anti- sheep IgG antibody (1 in 10, 000 dilution, Pierce) followed by enhanced chemiluminescence (ECL, Amersham).