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APOPTOSISKITS - M30‑APOPTOSENSE® ELISA
SCREENING FOR PRO-APOPTOTIC AGENTS
The M30‑Apoptosense®
assay is a versatile tool to for screen for pro‑apoptotic drugs (Hägg
et al. 2002).
In the FIGURE below, MDA‑MB‑231 cells (cytokeratin 18 positive human breast carcinoma) were seeded in 96 well microtiter plates at a density of 10 000 cells/well (in 200 µl medium). Treatment with the pro‑apoptotic candidates was initiated after 6 hours. After 40 hours, NP‑40 was added to 0,5% final concentration to the medium and the plates were frozen. By this procedure, it was possible to assay total caspase‑cleaved CK18 produced (cell associated + antigen released into the medium). 25 µl of the medium/extract was used for the M30‑Apoptosense® assay.
A number of pro‑apoptotic drugs were identified from a chemical library. The exact number of drugs that can be identified depends on the nature and size of the library used, the drug concentrations chosen and their time course kinetics.
3 non-treated controls were used
Substances to be tested were added to each well to a final concentration of 5µM.
Cisplatin was used at the following concentrations (left to right):
10 µM, 20 µM, 20 µM, 30 µM and 30 µM.
Results from testing 88 drug candidates for apoptotic activity.
Significantly higher pro-apoptotic activity was found in about 2% of the compounds screened so far.
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