星形孢菌素

從 Streptomyces staurospores 中分離出的 Staurosporine 是一種蛋白激酶抑製劑。Staurosporine 在 2-3 小時內以 1µM 誘導 DNA 片段化和細胞凋亡。

Background

Staurosporine is a natural product derived alkaloid which is isolated from Streptomyces staurospores cultures. This protein kinase inhibitor belongs to a family of kinase inhibitors containing an indole carbozol chromophore and is a potent inhibitor of a number of kinases. The ATP binding site on these kinase enzymes appears to be targeted by these Streptomyces derived inhibitors. Inhibition of these intracellular kinases leads to the induction of apoptosis as exhibited by the classic chromatin condensation leading to the formation of micronuclear bodies and reduced cell volumes. DNA is subsequently cleaved into oligonucleosomal fragments (laddering) as evidenced by agarose gel analysis. Staurosporine can be used with ICT’s apoptosis and cytotoxicity detection assays to create positive controls.

Reagent Name

Staurosporine

Target

Kinases

Sample Type

Cell culture

Storage

-20°C

Country of Origin

United States

Protocols

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.

Frequently Asked Questions

Question: I already purchased your FAM-FLICA kit for flow staining. I am trying to use Staurosporine to induce apoptosis as a positive control. Since the cells I am using are not cell line in culture, they are actually flushed out of mouse lungs. How should I use Staurosporine to induce apoptosis? Any method and advice to help? Answer: Staurosporine is a potent protein kinase inhibitor and is very effective at inducing apoptosis across a range of cell lines and types. I understand you are not working with a particular cell line, but instead are culturing cells after they have been flushed out of mouse lungs. I would suggest setting up an experiment where you evaluate a range of staurosporine concentrations and exposure periods. As a starting point, we normally use 1 µM concentration for 4 hours to induce apoptosis in >90% of Jurkat cells. We have found that some cell lines, such as U-937 cells, may require up to 6 hours for similar results. Thus it is best to set up a titration experiment with your cells and derive the optimal staurosporine concentration and exposure time experimentally.