Splitting Adherent Cell Lines
Carolina Lopez
Published: 2024-03-21 DOI: 10.17504/protocols.io.x54v9p51qg3e/v1
Disclaimer
Timing for trypsin treatment and following splits may need to be adjusted based on the different cell types considering how adhesive they are and their growth rate.
Abstract
This protocol describes how to split and maintain adherent cell lines in culture. Examples of these cells are: A549 cells, LLCMK2 cells, and MDCK cells.
Steps
Protocol
1.
Splitting Adherent cells
- Wash flask 2x with sterile PBS.
- Add 2mL of trypsin/T75. Incubate at 37 oC for 2-3 mins or until cells are detached from the flask.
- Add 8mL of cell culture media and pipette up and down. Transfer all the media to a 15mL tube.
- Centrifuge at 1200 rpm for 5 min.
- Add 2ml of cell suspension to T75 flask, then add 10ml Tissue Culture Medium. The cell will be ready for next split two days later.
