MEFs dying after passage - (Jan/19/2012 )
My problem looks like this:
On day 1, I thaw the cells, then plate them on Medium, after they reach confluency, usually on Day 2 I Passage the cells and then majority of them dies or doesn´t attach.
We´ve already used newly prepared Medium, Tripsine and PBS and still no change.. Also sometimes there is a yellow body-like thing in some cultures, but I´ve been told that this is a part of medium, the structures are bigger than cells and once some cells got yellow as well....
Does anyone have any idea what might be happening?
Thank you a lot for trying!!!
Are these primary MEFs or immortilized? Is it possible you are not quenching the Trypsin? Did you try adding new FBS? I don't know what you mean by yellow body-like thing....why don't you filter your medium to be sure it is sterile?
A picture of the yellow body would be helpful.
What passage are the MEFs if they are not immortalized?