1. Calculations II

2. Working With Molarity (M) In Some Cases Molarity (M) Is Known. Ex. 10 -5 M Compound X (MW 325) You Also Know That Cultures Should Be “fed” 30 ng/mL Of Compound X What Volume Of Compound X Stock Solution Should You Use To “feed” 10 mL Of Culture?

3. One Approach Is To Convert 30 ng/mL of cpd X Into Molarity Working With Molarity (M)

4. Alternative Approach Working With Molarity (M)

5. Compound Y: 3x10 -5 M, (MW 425), “Feed” 5 mL Culture with 10 ng/mL Ans: 3.9  L Exercise

6. If volume added is significant Ex. 1 mL, and Culture Volume is 10 mL First determine V 1 Based On Culture Volume (V 2 =10 mL) Re-determine V 1 Based on New Culture Volume (10mL+V 1 +0.5mL extra) When High Accuracy Is Needed

8. Determine what volume to use from stock vial –Culture volume is 11 ml –Stock vial concentration is 1mM –“Feeding” concentration is 0.1 mM –Final Culture Volume Should Be Between 12.25-12.5 mL Exercise

9. Solution

10. Cell Suspension of 2 mL @ 5x10 6 cells/mL Use a 24 well plate, Prepare 12 wells @ 0.25x10 6 cells/mL, 1 mL in each well Cell Calculations  Add 50  L To Each Well  Add 950  L Medium

11. Cell Suspension of 10 mL @ 0.25x10 6 cells/mL Use a 25cm 2 flask, Prepare 5 mL @ 1x10 6 cells/mL You will have to resuspend cells in smaller volume Even then, not enough cells for 5 mL @ 1x10 6 cells/mL Enough for 2.5 mL @ 1x10 6 cells/mL Cell Calculations

12. Prepare 12 wells @ 0.25x10 6 cells/mL Well volume: 0.5 mL Cell Suspension concentration: 10x10 6 cells/mL, 2 mL Answer: 13  L+487  L How Many wells can you prepare? Answer: (20x10 6 /0.125x10 6 ) =160 wells Exercises