Protocol for Raw 264.7 Cell Thawing

Protocol for Raw 264.7 Cell Thawing

1.     Warm water bath to 37 °C.

2.     Place 10 mL media DMEM in a sterile 15 mL centrifuge tube.

3.     Make your growth media (DMEM) for your new culture (10 ml DMEM in a 75 cm² Tissue culture flask)

4.     Take your cells out of nitrogen storage and thaw rapidly by swirling in the 37 °C water bath.

5.     Sterilize the outside of the vial with 70% ethanol, bring in to the culture hood and add the cells slowly to the media you prepared.

6.     Centrifuge at 1200 rpm, room temperature for 5 minute.

7.     Pipet off the supernatant. Then, add DMEM and pipet up and down to resuspend them in DMEM.

8.     Pipet the cells into a 75 cm² Tissue culture flask containing 10 ml DMEM and incubate at 37⁰C, 5% CO2 in an incubator for growth of your new culture.

9.     After overnight incubation pipet off the old media and add new media. The incubate again at 37⁰C, 5% CO2 in an incubator.