Hello everybody!
I'm having troubles with a previous established protocol for cells coculture in collagen on transwell for a 3D in vitro skin model. I prepared the collagen mix with 0.5M NaOH, 0.2M Hepes, 10× DMEM and fibroblasts, I poured this in transwells with pores of 0.4um and I let it polymerize for 60 min. After that I seeded epidermal cells on the top of the collagen and I put cell medium under the transwell. The day after the collagen scaffold was ok and I added on the top of it 500ul of medium. Two days later the scaffold was completely shrunk.
Has anybody ever had this problem??
Thank you
I was reading another paper on similar line and trying to compare your protocol of preparation of collagen gel. To me it looks you 0.5M NaOH may be too much, have you used this protocol before and have worked well before. if not than try to modify based on literature, good luck. I have seen some (in few wells) of shrinking of my cartilages in another system when I used to differentiate my stem cells to cartilage on collagen coated wells, though slight different system.
Thank you very much!! Actually I have used this protocol many times with 0.5M NaOH and this shrinking never happened to me before. So I don't know how to avoid it..
Dear Simona Villata,
Have you change any of current parameters especially cells number? before you faced the problem currently? You may try to prolong the incubation time for polymerisation or maybe seed the epithelial cells at different days...from my experience collagen hydrogel will be shrinking following time upon its maturity, the different is how severe the shrinkage effect..Best of luck!
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