I am trying to make a calibration curve using a 20mM stock compound in DMSO.
I want to achieve the following concentrations in this order, going down the plate:
I am setting this calibration curve on a Janus liquid handler. I have worked out how much compound to use in order to achieve the concentrations (i want to use as little volume of compound as possible as we are not supplied with a large volume). I also want each well to contain 5% DMSO and 20% acetonitrile.
1. aspirate 15μL of compound (20mM) into 285μL of 20% MeCN. This achieves a 1000μM conc. and 5% DMSO concentration.
2. Aspirate 5μL of compound (20mM) into well 2 which contains 120μL of 20% MeCN. This makes a conc. of 800μM and a 4% DMSO content if I am correct...
3. Aspirate 150μL of the 1000μM solution and dispense into 100μL of 5%DMSO;20%MeCN. This will give a concentration of 600μM but I am unsure as to what the overall DMSO content is now? Please could you help/provide workings out.
4. Aspirate 125μL of the 600μM solution and dispense into a well containing 125μL 5%DMSO;20%MeCN. This give a 300μM solution. Again I am unsure what the % DMSO content would?
5. A 2 fold dilution now occurs as described in step 4 until a conc. of 2.3μM is achieved in the 11th well in the plate.
Please could someone help me work out the % of DMSO that would be in each well? I have been going round in circles for weeks now. TIA.