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02/15/13
- HEK293-luc cell culturing, trial 2
HEK293-luc #4 cell culturing, trial 2
- Tried plating at the optimal density (1600 cells/well) in a 6-well plate; cells did not grow very well
- Retry 6-well plates and also try 96-well plate
- Made three new plates of HEK293-luc #4 cells
- Started with ~1x106 harvested cells (confluent T25 flask)
- Made the following dilutions:
- (1) 1:25 dilution of harvested cells (1mL cells + 24 mL medium) = 40,000 cells/mL 10% FBS DMEM
- (2) 1:25 dilution of previous tube (1mL 1:25 cells + 24 mL medium) = 1:625 dilution = 1,600 cells/mL 10% FBS DMEM
- (3) 1:100 dilution of harvested cells (100 μL cells + 9.9 mL medium)
--> 1:100 dilution of these (100 μL cells + 9.9 mL 20% FBS DMEM)
--> 1:4 dilution of these (5 mL cells + 15 mL 20% FBS DMEM)
= 25 cells/mL = 5 cells/200 μL (20 mL total)
Plates:
- 6-well plate: 2 mL of 1:25 dln., ~80,000 cells/ well
- 6-well plate: 2 mL of 1:625 dln., ~3,200 cells/ well
- 96-well plate: 200 uL of 1:40,000 dln., ~5 cells/ well (Note: was careful to gently mix stock tube by inverting after every column was filled to ensure that cells stayed suspended in the stock tube)
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