Coating Dishes with Gelatin - (Feb/28/2013 )

Does anyone have a protocol for coating cell culture dishes with 0.1% and 2% Gelatin? Also, do you use H2O or PBS to dissolve the gelatin? I am confused by the variation in protocols about the coating time and temperature:

(Protocols I have seen)
-Coat for 30 mins at room temp. then aspirate and use
-Coat 30 mins at 37C then aspirate and dry for 4 hours then use
-Coat, aspirate, then dry for 2 hours, then use
-Coat, 4C overnight, aspirate then use

Can someone tell me what works best for them? Thanks!

All of them work fine, choose the one that suits you. Having said that, for your first one I would extend the time out a bit - probably an hour or more would be better.

From my observations, coating time doesn't change anything.
However, some cells might not adhere if the gelatin isn't dry.

Thanks. Can't believe it can vary so much. But I guess the end result is to just have gelatin on the bottom of the container.

Hello all,

I would like to revive this question. I have not been able to pin down a standard protocol either. From the posts it looks like most any old method regarding timeline will work as long as it sits for an hour and is dried.

My interest is in autoclaving the gel. I have read a variety of protocols that autoclave at a variety of temperatures for a variety of times. When I autoclave and place the .1% gelatin in ddH20 in the fridge it never firms up, even after only 15 min at 121C. Whereas if I don't autoclave the solution will become firm in the fridge.

Any thoughts on autoclaving a .1% type A porcine gelatin solution?

Thank you for your time! Hopefully I can contribute to this great community!

I've just been putting the 0.1% gelatin (autoclaved) coated dish in the incubator (37C) for 30 min, aspirate, then use. Results are fine so far. You let your surface dry before seeding?

To address your question about the gelatin not firming up, I'm not sure why that is the case. Perhaps an enlightened one will shine their light upon this. Only my 2% gelatin firms up at 4C.

Thanks for the response jeee!

I do make sure I let the surface dry before using it. I am growing tail tip fibroblasts and they grow alright using my current methods. I thought I might be able to get some faster/better growth if I cleaned up my protocol.

I am concerned that the gel isn't firming up at 4C. Thinking that the gel might be breaking down due to the autoclave.