Is 3T3-F442A a clone of NIH-3T3 or FAA2A transfected 3T3? sometimes, clone are very different from orjinal cell line for example P2X7 transfected HEK-293 cells are very different from HEK-293 cells in terms of attachment properties, growing rate, morphology etc. So,

1) your problem may depend on the clone.

2) we use corning high-binding culture plates for the clones easily detach from plate surface. you can try that plate.

3) you could centrifuge the cells after detachment of the cells with trypsin and use new flask each passage. cells grow properly new and fresh media and places (tebdil-i mekanda ferahlık vardır :)

differentiation is pretty strong on cells, they coul detach because they are stressed or suffer the differentiation stimuli. if the detached cells are dead, could be that are suffering so you should modify your differentiatio protcok, if the cells are alive, you could coat the plate to favour adhesion, like with poly-lyisn (L or D) for 30min-2h in the incubator, than wash and pate your cells, good luck!

Thank you for your answers. This clone is widely used in the literature and no one mentioned about this problem. A common protocol used is starting differentiation 2 days after confluency. However, we were not able to start differentiation process as cells detached as a layer 2 days after confluency. We obtained a new vial and now we have the same problem. We did not tried high binding plates or coating methods and those would probably work. However, none of the articles using this cell line mentioned about these approaches. They used standard culture plates and standard culture techniques. So there must be something wrong. I want to figure out this problem.

Presently, I´m working with cells close to 3T3-F442A, the cell line 3T3-L1.  I attach a very good techical paper for differentiation these cell line successfully.  Good Luck¡

unfortunately, you can't find any problem about the experiments in articles. The authors write the article as if everything is OK and "standard". My HEK-P2X7 cells were terrible and didn' t attach. I tried every methods for them but i couldn't find any report about this problem until i met the researchers in a conference.

I have recently purchased the same cells from sigma and I also experience cell detachment after induction of differentiation. I have worked with this cell line before and never had that issue. I also never had to coat my plates before. Perhaps it is due to this particular batch of cells? 

The differentiation into adipocytes depends on the maintenance of contact inhibition and it has reported that the 3T3-F442A cells have lost their contact inhibition potential (see info on their site and on that of the ATCC/ECACC). The problem is also present is the 3T3-L1 cells, so they are no  alternative. 

What is the cell density you use when plating cells prior to the differentiation assay? Do you plate them in low density and allow them to become confluent over the next few days, or do you plate them at high density to obtain confluence the next day?

Dear Matthias,

I have worked with 3T3-L1 cells obtained from ATCC and there was no problem in differentiation. Once, we had seen the same problem for 3T3-L1 cells and realized that we overconfluenced the cells during propogation and ordered a new vial. The problem was solved. 

When we first observed this problem in 3T3-F442A cells, we ordered a new vial as we thought we somehow could overconfluenced the cells during expansion. Although we were extremly carefull in this vial, the problem still existed.

We plated 1x10^4 cells/cm^2 and waited two days to allow cells to become confluent. From this point, cells started to detach (Please see the attachment)

Use Cell Bind plates from Corning. Additional precoat with 0.1% collagen may also be beneficial

Hi, we regularly work with 3T3F442A cells, this cells don't have any trouble if the culture dish is 60mm or above, for smaller culture dishes or even slides, you need to coat the surface with poly-lysine, to help the cells to hold to the surface.