Killing+Cells+with+Camptothecin+for+Positive+Control

Original Protocol:

//Induction of Apoptosis by Camptothecin// >>>>> 2 uM -> 2 uL >>>>> 6 uM -> 6 uL
 * Prepare a 1.0 mM stock solution of camptothecin (Sigma-Aldrich Cat. No. C-9911) in DMSO (maybe 1% DMSO is better since this would harm the cells less and most likely still dissolve the camptothecin). Camptothecin, an extract of the Chinese tree Camptotheca acuminata, is a potent inhibitor of topoisomerase I, a molecule required for DNA synthesis. Camptothecin has been reported to induce apoptosis in a dose dependent manner in vitro.
 * Camptothecin molecular weight: 348.4 g/mol
 * 1 mmol/(1000 mL)* 1 mol/ (1000 mmol) * 348.4 g/(1 mol) * 1 mL = 3.484*10^(-4)g or 0.348 mg
 * Add camptothecin to 1X10e6/mL proliferating Cells. We will probably use the max concentration of cells that we can fit in 3 mL. Concentration of cells measured using trypan blue.
 * Groups
 * No camptothecin control
 * 0.1 uM
 * 2 uM
 * 6 uM
 * Jurkat (6 uM) (maybe. . . these cells weren't growing well.
 * Volume 1 mM stock needed
 * 1mM*y/(1000*uL)=0.0001 mM -> y=0.0001 mM * 1000 uL / (1 mM)
 * 0.1 uM -> 0.1 uL (of 1 mM into 1 mL)
 * Incubate the cells for at 37 C
 * The recommended time is 4 hr. However, the incubation time will depend on the cell type.
 * We were able to detect killing of Jurkat cells with a 4 hr incubation, but no killing of NIH 3T3 fibroblast cells was detected with this time. The company recommended using a 24 hour incubation.