culturing MCF-7 cellls - clumping and maintaining healthy cells (Feb/15/2010 )

hi! It's my first time to handle cell culture and I don't really have much background about it. I just wanna ask if how can i maintain a healthy culture of MCF-7 cells? How do I know if my cells are still healthy or not? dead or alive? can you show me pictures of healthy, unhealthy, and dead cells? because my MCF-7 cultures are mostly round and small, like they have shrunk a bit, and not like what i can see in the ATCC photo, which is elongated. many also have lots of dots in them. moreover, my cells are often very dense and there are so many clumping. I regularly washing them or feed them every two days when they are not yet confluent to remove floating cells and to lessen the clumping, yet they are still clumping so much I cannot make a clean monolayer of cells. please help me.

thanks!

PS. attached is a photo of my MCF-7 cells.. sorry if its quite unclear.. i was using my own camera...

What you have are some cells that need to be split/passaged again... the clumping is probably due to not making the cells into a single cell suspension when plating/seeding. This may be because you have under trypsinised them;thereby not letting them separate enough so that when you pipette to break them up, they don't separate. Over trypsinising will also cause this to happen, the cells will cling to each-other.

You also need to make sure that you are not seeding at too high a density - anything over 50% of the area covered will mean you will probably have to split them again within a day or so.

I suggest that you get someone experienced to sit beside as they do their culture so you can see their techniques. Also check out a book on cell culture such as R.I. Freshney, Culture of Animal Cells: a laboratory manual.

Your MCF-7 cells look normal. I regularly culture MCF-7 cells and they do clump. If you look at the picture from ATCC again: http://atcc.org/Attachments/1980.jpg, you will notice the clumping of the cells. I asked ATCC last year about this since I also thought that my MCF-7 cells are unhealthy since they clump. Here is ATCC's reply: "We noticed in the most recent lot that these cells got to be 60-70% confluent in 2-3 days in a T-25 (10mls), but took another 5 days to reach a monolayer (about a week total). They grow in clusters/patches, and you will even notice viable, floating cells. Please keep in mind that we recommend subculturing the cells between 70-80% confluency."

Just make sure you use the right media (do not forget the insulin!). Hope this helps.

Hi! everybody! i'm culturing MCF-7 cell line and i want to isolate breast cancer stem cell from that. But i don't know how to do. Please help me! Thanks!