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detach RAW264.7 - Who knows an enzyme to detach RAW264.7??? (Jul/01/2009 )

Hi everybody,

Im new to cell culture and Im working with RAW264.7 (mouse macrophages).
I have to do a viability count with trypan blue and for this I need to detach the cells from a 6 well-plate.
I scrape them off with a cellscraper at the moment but I have got the feeling this is not good for them because their vitality is much lower than that of my other cell lines (that are detached enzymatically).

Trypsin or trypsin/EDTA do not work with RAW264.7 cells. Scraping is the suggested method by the supplier but perhaps someone has developed another way?!

Thanks in advance,

Vrieschen

-Vrieschen-

Vrieschen on Jul 1 2009, 09:59 AM said:

Hi everybody,

Im new to cell culture and Im working with RAW264.7 (mouse macrophages).
I have to do a viability count with trypan blue and for this I need to detach the cells from a 6 well-plate.
I scrape them off with a cellscraper at the moment but I have got the feeling this is not good for them because their vitality is much lower than that of my other cell lines (that are detached enzymatically).

Trypsin or trypsin/EDTA do not work with RAW264.7 cells. Scraping is the suggested method by the supplier but perhaps someone has developed another way?!

Thanks in advance,

Vrieschen



We grow these cells in suspension and therefore it is easy to do a viability count. If you then do an experiment in 6 well plates and want to do viability, then you will have to do them in-situ i.e. do not scrape or try to trypsinise, leave the cells attached, add the trypan, use a graticule and count.

RAW and J774 cells stick to plastic like superglue...impossible to trypsinise and grow like most other cells. That's why we grow them in suspension.....viability 99-100%

Hope this is useful

Kindest regards

Uncle Rhombus

-rhombus-

We grow these cells (and all macs) on petri dishes. Since the plastic is not treated to increase cellular adherance, it is much easier to remove the cells. These little buggers practically grow into tissue-culture plates!

-miBunny-

Hi Rhombus,
How do you culture these cells in suspension? Is there a manual that I can refer to?
Thanks for your help!

-cmubio2010-