Jurkat cells are cultured in RPMI medium supplement with 10% FBS. The cells were grown to late growing phase. Anisomycin is added to the medium to a final concentration 10 µg/ml (BioVision cat# 1549-10, dissolved in DMSO to 10 mg/ml as stock). The cells are incubated at 37°C for 15 minutes, then collected and lysed in Extraction Buffer (BioVision Cat # K430-40). The cell lysate can be used as a positive control for JNK activity assays in conjunction with BioVision’s JNK Activity Assay Kit (K430-40 or K431-40). For positive controls, we recommend using 100 µg per assay.