Sean Lauber:Using the Lewis Lung Carcinoma cell Mouse Model for Lung Cancer: Difference between revisions
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6. Plate this volume on a single 100 mm non-tissue culture treated petri dish | 6. Plate this volume on a single 100 mm non-tissue culture treated petri dish | ||
7. Place at 37*C (with 5% CO2) for 72 h | 7. Place at 37*C (with 5% CO2) for 72 h | ||
8. Recover the cells by pipetting and then ejecting the media onto the plate | 8. Recover the cells by pipetting and then ejecting the media onto the plate | ||
* repeat this twice and do it VERY GENTLY to ensure you don't disturb the clumps | |||
9. Split 1:3 (put 1/3 volume back onto original plate, and the other 1/3 onto two new plates) | 9. Split 1:3 (put 1/3 volume back onto original plate, and the other 1/3 onto two new plates) | ||
10. Bring the volume up to 10 ml with 10% FBS DMEM | 10. Bring the volume up to 10 ml with 10% FBS DMEM | ||
11. Place at 37*C (with 5% CO2) for 24 h | 11. Place at 37*C (with 5% CO2) for 24 h | ||
12. Collect cells by gently pipetti | 12. Collect cells by gently pipetti | ||
Revision as of 12:36, 13 August 2012
For 5 animals:
1. Thaw a single vial of 1,000,000 cells 2. Add to 9 ml of warmed 10% FBS DMEM 3. Spin at 10,000 RPM for 3 min 4. Discard supernatant 5. Resuspend to 10 ml 6. Plate this volume on a single 100 mm non-tissue culture treated petri dish 7. Place at 37*C (with 5% CO2) for 72 h 8. Recover the cells by pipetting and then ejecting the media onto the plate * repeat this twice and do it VERY GENTLY to ensure you don't disturb the clumps 9. Split 1:3 (put 1/3 volume back onto original plate, and the other 1/3 onto two new plates) 10. Bring the volume up to 10 ml with 10% FBS DMEM 11. Place at 37*C (with 5% CO2) for 24 h 12. Collect cells by gently pipetti
