Altman:Protocols/TC Protocols/Freezing SF9: Difference between revisions
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6. Resuspend the cells in the media+DMSO to return the cells to a final density of greater than or equal to 1*10^7 viable cells/mL | 6. Resuspend the cells in the media+DMSO to return the cells to a final density of greater than or equal to 1*10^7 viable cells/mL | ||
7. Aliquot 1.5 mL of cells into 2.0 mL cryovials | 7. Aliquot 1.5 mL of cells into 2.0 mL cryovials | ||
Latest revision as of 13:15, 24 July 2013
Department of Physics, Willamette University
Freezing down SF9 cells for long-term storage
1. Grow the cells to a density that is greater than or equal to 1*10^7 viable cells/mL
2. Pellet the cells by centrifugation: place the 35 mL of cells in a 50-mL conical, and spin at 1000 rpm for 10 minutes
3. Remove the media; this media is "conditioned media"
4. Combine the conditioned media 1:1 with fresh media
5. Add DMSO to the media to a final concentration of 7.5% (e.g. 2.6 mL DMSO + 35 mL media)
6. Resuspend the cells in the media+DMSO to return the cells to a final density of greater than or equal to 1*10^7 viable cells/mL
7. Aliquot 1.5 mL of cells into 2.0 mL cryovials
8. Place tubes between two pieces of styrofoam and place in a -20 freezer overnight
9. Place in liquid nitrogen for long term storage
