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Monday
HCT cells from the previous week were split into six different 10 cm square plates. The plates were labelled as HCT +/+ 1D; HCT +/+ 2D; HCT +/+ N; HCT -/- 1D; HCT -/- 2D and HCT -/- N. All cells are at passage three. In addition two T25 flasks containing HCT +/+ P2 and HCT -/- P2 were frozen down and stored into -20 degrees celcius freezer. -
Period: to
FYP
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Tuesday
All six plates were checked for cell growth and contamination. Everything is good. *Incubator was cleaned and four flasks were discarded due to bacteria contamination. -
Wednesday
Change media of all 1D and 2D plates to serum free media. HCT -/- N may require splitting to maintain a appropriate cell density. Frozen cells from Monday was transferred from -20 to -80 freezer. -
Thursday
Collect supernatant (media) from HCT +/+ 1D and HCT -/- 1D. Store in Falcon tube and spin down to remove dead cells. Transfer spun down supernatant to new tube and keep at -20 degree celcius freezer. DO NOT DISCARD SUPERNATANT AFTER CENTRIFUGATION. Change medium of HCT +/+ N. -
Friday
Repeat collection procedure for all 2D plates. Split both HCT +/+ N and HCT -/- N with a 1:10 ratio using NORMAL medium.