We have obtained 3T3-L1 from ATCC and we started propagating cells in DMEM-High glucose with 10% FBS. We followed the standard protocol for differentiation of cells that is., seeding cells in 24 well plate (20,000 cells/well/ml). Day 2 of post confluence, we started differentiating cells using Dexamethasone, IBMX and insulin. initially when changing the medium there was no curling up of cells. But after cells differentiate, when we change the medium the cells from edges are curling up thereby our experiment is abruptly terminated because of curling up of cells. I tried manual changing of medium slowly dropwise using micropipette. We even tried different types of serum like, FBS and Calf serum which was not of great help.
Can anyone suggest us a solution to solve the problem of curling up of cells when changing medium..

that will/might come a little too late, I have only seen this post now...

1.) be sure that the medium is pre-heated to 37°C
2.) what you could try is 'conditioning' the medium, i.e. putting it on a large* confluent plate of undifferentiated cells for - let's say - 24 hours, before giving it to the cells for the assay
but I am not sure how stable insulin is....

*...so that you have enough of it

hope you have solved the problem by now

you also could do only a partial medium change (50%?) if that works for the assay

Did you solve the problem of curling up of your 3t3-L1 cells after differentiation?

Hi did u get any solution for 3t3l1 curling up cuz even i am facing the same...........

Were there any other solutions for this problem? I have had the same issue for 3 separate lots of cells...

Hi - It seems like alot of people - myself included - are having the same problem with curling up of cells during differentiation. The conclusion of a previous thread ("3T3-L1 issues") was that it looks like the latest lots of 3T3-L1 cells from ATCC are not good and to get older lots of cells from people who are happy with them. I guess I will try this. By the way, the "3T3-L1 issues" thread started in 2006. I'm not sure if that means that all the 3T3-L1 lots from ATCC starting from back in 2006 have this curling up problem....

Also see the other 3T3 post - you can use the 'search site' option on the Solutions Search bar on the left. That covered a whole gamut of other possible causes including mycoplasma, medium components etc.