Passaging C2C12 cells

From Tanaka Wiki
Revision as of 08:45, 28 April 2014 by Stephanh (talk | contribs)
Jump to navigation Jump to search

Materials:

  • 1x 15ml FALCON tube
  • PBS w/o Ca2+ and Mg2+
  • TE in PBS (4 ml TE + 36 ml PBS)
  • HS for C2C12 cells
  • pipettes
  • 75 cm2 flask

procedure:

  • remove 5 ml of the old media in the 15 ml FALCON tube
  • throw the rest of the old media away
  • wash the cells with 5 ml PBS
  • remove the PBS
  • add 2 ml of the TE in the dish
  • let this incubate for ~4min at 37ºC
  • stop the TE with the old media and resuspend the cells in this
  • pipette the cell suspension in the FALCON tube back
  • centrifuge the suspension for 1000 xg for 3 min at 4 ºC
  • remove the old media
  • resuspend the cells in an appropriate volume of HS medium (f.e.: 10ml)
  • fill in the new 75 cm2 flask 14 ml of new media
  • pipette an appropriate volume of the cell suspension in he new flask and resuspend it in the new flask (f.e.: if you want to dilute your cell 1: 20 put in the new flask 0.5 ml of the resuspended cells in the FALCON tube)
  • close the flask and put it in the 37 ºC