Camptothecin

1. Dissolve staurosporine powder in tissue culture grade DMSO to obtain a 1 mM staurosporine concentration., 2. Prepare 50 – 100 µL aliquots of the DMSO solubilized staurosporine stock solution and store them frozen at < -20°C. A frozen vial of staurosporine may only be rethawed 2X before it must be discarded., 3. Spike cell cultures at a staurosporine concentration of 1 µM in the cell culture media. This equates to a 1 µL spike of the 1 mM staurosporine stock per mL of the cell culture suspension. This concentration works Well for inducing many cell lines including HL-60 and Jurkat cells when using a 4-5 hour 37°C incubation period. Typical cell suspension concentrations that have been used for this staurosporine induction protocol range from 1 x 105 – 1 x 106 cells/mL., 4. Perform time course studies on your particular cell line to ascertain the optimal staurosporine concentration and exposure time required to achieve good apoptosis induction levels in your experimental system., 5. Proceed with your experimental apoptosis induction model system.

Camptothecin is a potent cytotoxic pentacyclic alkaloid known for its potent apoptosis inducing properties. Camptothecin and its analogs have been demonstrated to inhibit topoisomerase I (topo I), causing cells in S phase to enter into apoptosis. Camptothecin binds to and stabilizes the topo I – DNA backbone complex (covalent binary complex). This stabilized complex diminishes the rate of topo I release from the broken strand, thus slowing the religation and subsequent DNA synthesis process. Use camptothecin to create positive controls in apoptosis detection experiments.