Western blot analysis of extracts from A172 cells, untreated or treated with indicated concentrations of Bisindolylmaleimide I for 30 minutes, followed by stimulation with 200 nM TPA for 10 minutes. The phosphorylation of PEA-15 was detected by using Phospho-PEA-15 (Ser104) Antibody #2776 (upper). PEA-15 Antibody #2780 was used as a loading control (lower). Bisindolylmaleimide I-mediated inhibition of TPA-induced PKC phosphorylation of PEA-15 at Ser104 is shown.
Bisindolylmaleimide I structure
Bisindolylmaleimide I is supplied as 500 μg powder. Store at or below -20ºC. Before use, dissolve powder in 0.28 ml DMSO to make a 4 mM bisindolylmaleimide I stock solution. For working concentrations of 2 μM-4 μM, dilute DMSO stock 1:2000 to 1:1000. Treat cells with the desired concentration for 30 minutes.
Store lyophilized or in solution at -20ºC, desiccated. Protect from light. In lyophilized form, the chemical is stable for 24 months. Once in solution, use within 3 months to prevent loss of potency. Aliquot to avoid multiple freeze/thaw cycles.
C25H24N4O2 • HCl
Bisindolylmaleimide I (BIS) is a potent inhibitor of PKC (1,2). In vitro the IC50 of BIS is 10-20 nM for PKCα/β/γ and 100-200 nM for PKCδ/ε isoforms. The in vitro IC50 for PKCζ is about 6 μM, indicating that BIS is a very weak inhibitor for this isoform. In in vivo cellular assays the IC50 of BIS for PKC is between 0.2-2 μM (1,3).
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