User:Wilfred J. Poppinga/Notebook/cAMP compartmentalization/2010/03/26: Difference between revisions

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===Plating cells from -80 °C===
===Plating cells from -80 °C===
*Take a Ø10cm dish.
*Take a Ø10cm dish.
*Put 10 ml of medium in a 15 ml Falcon tube.
*Put 10 mL of medium in a 15 mL Falcon tube.
*Get the cells out of the -80°C.
*Get the cells out of the -80°C.
*Defrost the cells quikly by using a water bath.
*Defrost the cells quickly by using a water bath.
*As soon as the cells are defrosted, pipet the cells into the medium which is in the Falcon tube.
*As soon as the cells are defrosted, pipette the cells into the medium which is in the Falcon tube.
*Put the medium containing the cells into the petridish.
*Put the medium containing the cells into the petridish.
*Put O.N. at 37°C
*Put O.N. at 37°C

Revision as of 08:36, 26 March 2010

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Summary

  • Plate cells for Anouk
  • Plate cells to bring to Berlin
  • PROTOCOL HEAT INACTIVATE FBS/FCS
  • PROTOCOL USING CELLS FROM -80°C

Materials & Methods

Plating cells from -80 °C

  • Take a Ø10cm dish.
  • Put 10 mL of medium in a 15 mL Falcon tube.
  • Get the cells out of the -80°C.
  • Defrost the cells quickly by using a water bath.
  • As soon as the cells are defrosted, pipette the cells into the medium which is in the Falcon tube.
  • Put the medium containing the cells into the petridish.
  • Put O.N. at 37°C
  • Look the next day if the cells are attached.
  • Wash the cells twice with PBS.
  • Incubate in fresh medium.
  • If the plate is now confluent the cells can be split and used for experiments.

Preparation of Heat inactivated FBS/FCS

  • Defrost FBS @ 4 °C (over the weekend)
  • Incubate exactly 30 min. @ 56 °C and put to ice
  • Prepare sterile aliquots
  • Store @ -20 °C


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