What can I do with my MCF7 cells? - (Dec/17/2013 )

Well I wanted to conduct my experiment using MCF7 cells. I had a plate that was ~ 90% confluent, so I split my cells yesterday into a 12-well plate, a 6-well plate & a 60mm plate (plus a 10-cm plate for backup).

For my 6-well plate, I needed it to be 90% confluent.

For my 12-well and 60mm plate they need to be ~ 60-70% confluent.

I was hoping to conduct my experiment today with my 12-well plate & here lies the problem.

When I looked at my 12-well plate it looked alright & I was convinced it would be ready today. Looking at it TODAY, I hardly see any cells!!! It barely looks 30% confluent!!! With my other plates they look fine, so what went wrong?

I was really hoping to start my experiment today & I really can't afford any delays.

The good news however, is that I have a spare 10-cm plate that's about 50% confluent. I just don't know if it would be better to wait a few days to see more cells, or can I split my cells again from my 10-cm plate and dispense some cells onto my 12-well plate (even though I had split my cells yesterday)?

Would someone be so kind as to help this noob out please?

Thank You.

Kenya McCalmont on Tue Dec 17 16:12:52 2013 said:

 

When I looked at my 12-well plate it looked alright & I was convinced it would be ready today. Looking at it TODAY, I hardly see any cells!!! It barely looks 30% confluent!!! With my other plates they look fine, so what went wrong?

 

I was really hoping to start my experiment today & I really can't afford any delays.

 

The good news however, is that I have a spare 10-cm plate that's about 50% confluent. I just don't know if it would be better to wait a few days to see more cells, or can I split my cells again from my 10-cm plate and dispense some cells onto my 12-well plate (even though I had split my cells yesterday)?

What do you mean that you looked at your plate and were convinced that it was alright?  Did you do this after the cell had settled and started to attach, or was it immediately after seeding? If it was immediately after seeding, then the only thing you can tell from that, is that you added some cells.  After they have settled there is a better chance of getting an estimate of how well you have seeded, but it is still highly subjective.  MCF7 tend to clump so area coverage is not ideal for estimating the confluency, you are perhaps better off seeding a certain number of cells that in other less clumpy cell lines would give you the confluency that you require.

I repeat the whole seeding procedure, adding more cells gives you two populations of cells that are different ages in terms of how recently they were seeded, which may have an effect on your results.