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Thank you for you're quick reply.
I used 3 different concentrations (1uM, 2 uM and 5 uM) for 4 hours. To detect apoptosis I used 2 different kits detecting activity of caspase 3. To detect cell death I used propidium iodide. To analyse the cells I used FACS analyser and the sturosporin treated Jurkat cells looked exactly the same as non treated cell control.
Once the sample arrived i brought it to room temperature and dissolved in 100 uM DMSO, prepared 10ul aliquots and stored them at -20*C. One of the aliquots (before freezing) I diluted with RPMI media to concentration of 200 uM and added to cell suspension. However once the vial with staurosporine arrived I could not really see the content- is the substance transparent?
Hope that tis information will help you to spot the problem.
I look forward to hear from you soon.
Asked on Apr 11 2012
Thank you for getting back to me and for providing some further details.
I have discussed your enquiry with my colleagues in the Lab. Could you please send details of the calculations used to reconstitute the solution.
We are a bit confused regarding the concentrations stated. It may just be a typo but it is very unlikely to produce 200 uM from a 100 uM stock solution.
We are also wondering if this compound has been used at a concentration that may not be right. However, we have found an article that suggests that a lower concentration of Staurosporine is 10-7 M and a high concentration is 10-6 M. These concentrations fall within the range you have stated.
Thank you for your understanding and co-operation in this matter. I look forward to hearing from you soon and resolving this issue as soon as possible.
Answered on Apr 11 2012