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Cell Culture Media, Kits, and Products |
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Subculturing Cells - Cell Culture
Cell Culture
Cell Subculture Protocol
SUBCULTURING CELLS
Get DMEM media 4C; Trypsin-20C; BSA-20C;
Put in 36mL of DMEM into conical tube plastic
Add 4mL of BSA media to 36mL DMEM;
Get syringe and filter;
Suck up media with syringe. Add filter.
Push out media through filter on syringe.
Repeat twice to get all of volume.
Put into new plastic tube conical.
Remove old media by dumping into beaker; DO NOT USE ASPIRATOR (get contamination!!)
Add 3mL typsin to culture. Trypsin with pull cells off the culture container.
Put container into incubator. Wait 2 min; Take out; look into microscope.
Try to get less clumps. More individual cells.
Remove most of trypsin by dumping into beaker.
Put back into incubator. Check frequently.
Slap container on sides to check bottom for less clumps; look into microscope.
When see many individual cells floating along; Great!
Add media to larger container; 1:5/ 1:3 dilution (3-5 containers larger).
Add 10mL to (T75)large container, 5mL to (T25)small container.
Suck out Most (5mL of cells from T25). Add to T75.
Leave some cells in small container. Add media to small container. Check in microscope for single cells; Incubate in 37C incubator.
See other Cell-protocols at our Cell Biology and Cell Culture Protocol Directory.
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