Problem with Caco-2 washing - cells detaching (Dec/06/2007 )
Hello,
I'm culturing Caco-2 cells in 25cm2 flasks, 24-well plates and cultureslides (all from BD Biosciences, standard TC). Many protocols require washing cells 1-2 times with PBS and that's where I'm encountering a problem - cells are detaching, monolayer is coming off the surface. I's happening almost regardless of the confluence. Anyone have and idea why it's happening?
Caco-2 cells are tricky. I dont like to work with them.
Did you use d-PBS with Ca2+,Mg2+ ?
Gibco/Invitrogen Ca/Mg free PBS in tablets.
try the one with Ca2+,Mg2+ and keep washing step brief.
I'm culturing Caco-2 cells in 25cm2 flasks, 24-well plates and cultureslides (all from BD Biosciences, standard TC). Many protocols require washing cells 1-2 times with PBS and that's where I'm encountering a problem - cells are detaching, monolayer is coming off the surface. I's happening almost regardless of the confluence. Anyone have and idea why it's happening?
Dear K.B.
This is a standard misconception that ALL tissue culture plastic is standard. This is definitely not the case. The BD products you are using I assume are Falcon TC plastic. This is the premier TC plastic and the leader in the field (UK Market).
BUT cells will grow differently on different TC plastics i.e. attachment, morphology and growth can be affected. Also the quality of the serum is important....what serum are you using?
Take home message, try different TC plastics:
Nunc, TPP, Sterillin, Costar, Triple Red, Griener etc etc etc.
Kindest regards.
Rhombus
It's really weird what is happening with your caco. Normally they are so difficult to detach from the plate. First of all I would try with another criostock of cells. Maybe your cells are too old.
Rigth, I rembered that I had to treat them for real long time with Tryp/EDTA to get them off, and the cells got damaged as a result.