Difference between revisions of "Splitting Cells"
From Bridges Lab Protocols
Davebridges (Talk | contribs) (added temperature) |
m (→Protocol: added the aspiration processes and changed wash cells twice to "once") |
||
| Line 6: | Line 6: | ||
==Protocol== | ==Protocol== | ||
#Warm PBS and Media in water bath | #Warm PBS and Media in water bath | ||
| − | #Wash cells | + | # Aspirate media |
| − | #Add 1 mL trypsin and sit in the hood for 2-5 min | + | #Wash cells once with 10 mL (per 10 cm dish) PBS -/- then aspirate the PBS |
| + | #Add 1 mL trypsin and allow to sit in the hood for 2-5 min | ||
#Add 10 mL media to each new dish | #Add 10 mL media to each new dish | ||
#Check cells for trypsinization, and if necessary tap the cells | #Check cells for trypsinization, and if necessary tap the cells | ||
Revision as of 17:28, 1 June 2017
Materials
- Media (FBS or NCS as required): Prepare by filtering together 5 mL PSG, 50 mL COS-FBS and one bottle of DMEM
- PBS -/-
- 0.05% Trypsin
Protocol
- Warm PBS and Media in water bath
- Aspirate media
- Wash cells once with 10 mL (per 10 cm dish) PBS -/- then aspirate the PBS
- Add 1 mL trypsin and allow to sit in the hood for 2-5 min
- Add 10 mL media to each new dish
- Check cells for trypsinization, and if necessary tap the cells
- Add 9 mL media to trypsinized cells
- Add 1 mL cells to each dish (for 10X dilution; 0.5 mL for 20X)
- Replace plates in the 37C incubator
Cell Specific Notes
- 3T3-L1 fibroblasts have special considerations regarding confluence. See Differentiation of 3T3-L1 Cells
- RAW 264.7 cells are scraped, not trypsinized. See Culturing RAW 264.7 Cells
- S2 cells are grown at 28C without extra CO2. See Culturing S2 Cells
