WinterVomitingLab:Protocols/RAW 264.7 cells
Return to WinterVomitingLab
Passaging RAW 264.7 Cells.
- RAW 264.7 cells (ATCC # TIB-71)
- Complete MNV DMEM
- DMEM (Fisher- cat# SH30022LS)
- 10% HyClone FetalClone III Serum (VWR- cat# 16777-240)
- 1% HEPES (VWR- cat# 12001-710)
- 1% Penicillin/Streptomycin (VWR- cat# 16777-164)
- 1% Sodium Pyruvate (VWR- cat# 45000-710)
- Determine the number of new flasks to be made, and label with passage #, cell line, date, and initials. Add the appropriate amount of fresh media for the size flask you are using (T25- 7ml, T75- 25ml, T175- 50ml), minus the amount you will be adding from the passaged flask. RAW cells are passaged at a ratio of 1:5 during the week and 1:10 over weekends.
- Pour media from each flask into the waste beaker, cover beaker, and set aside.
- Add 10ml of fresh media and, using a new cell scraper for each flask, gently scrape the cells from the bottom of the flask using a side to side motion, working from the bottom of the flask to the top. DO NOT move back down in the flask once you have gone up!
- Using a pipet, rinse the flask thoroughly to remove and mix all cells, then aliquot them into the appropriate number of new flasks containing fresh media.
- Discard old flasks and place new flasks in the 37° C incubator.
Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!
- List troubleshooting tips here.
- You can also link to FAQs/tips provided by other sources such as the manufacturer or other websites.
- Anecdotal observations that might be of use to others can also be posted here.
Please sign your name to your note by adding '''*~~~~''': to the beginning of your tip.
Relevant papers and books
Error fetching PMID 17133824:
Error fetching PMID 22951568:
- Error fetching PMID 15562321:
- Error fetching PMID 17133824:
- Error fetching PMID 22951568:
Video protocol produced by Katie Roache:
- Who has experience with this protocol?
or instead, discuss this protocol.