Dear Federica, just coat your flasks under the hood, let gelatin to polymerize on the incubator (37°C) for 1h and then, discard the excess under the hood.

Use parafilm to seal and store your flasks/plates in the fridge for a couple of days.

Before use, let the flask/plate to warm 10 minutes in the incubator.

To "test" the method, prepare one plate in this way and try ;)

I used it for gelatin and fibronectin coated cover slips and worked well...

Dear Federica, just coat your flasks under the hood, let gelatin to polymerize on the incubator (37°C) for 1h and then, discard the excess under the hood.

Use parafilm to seal and store your flasks/plates in the fridge for a couple of days.

Before use, let the flask/plate to warm 10 minutes in the incubator.

To "test" the method, prepare one plate in this way and try ;)

I used it for gelatin and fibronectin coated cover slips and worked well...

Dear Federica

1.Make up a 0.2% gelatin solution (0.2 gelatin powder in 100 ml sterilized DW)

2.Sterilize by autoclaving

3.Add 5 ml of the 0.2% gelatin solution per 35 mm sterilized dish under the hood

4.Leave for 1 hour under the hood.

5.Removed excess gelatin and let the dishes open in a laminar flow hood to dry (10-15 min).

6.Re-sterilize by UV under the hood for 30 minutes

7.Dishes are now ready for seeding and can be sealed by parafilm and stored in the fridge.

Best regards

Dear Federica

Agreed that you would need gelatin to coat for attachment and differentiation of EBs but why would you require gelatin for mesenchymal stem cells?

Isn't the property of MSCs to attach to plastic?

I think you should not have a problem if you are using serum in your media for culturing MSCs because some proteins in the serum help the adhesion of MSCs to plastic.But yes if you are using serum free media,then you will need a coating of any ECM component or maybe gelatin.

Nevertheless yes you can coat your flasks with gelatin(0.2%) and keep them in the incubator for 30 minutes.Then wrap the flask in parafilm and store it in 4 degrees for 3 days.

With best regards!

Dear Federica

I use gelatinated culture plate for differentiation EB TO HEPATOCYTE

Make a 0.1% gelatin solution and Sterilize by autoclaving then coat culture plate with adequate gelatin solution and put the plate in the incubator for 1 hour.Removed excess gelatin and let the plate open in a laminar flow hood to dry .after these work you can wrap the flask in parafilm and store your gelatineted culture plate in 4 degrees .

Dear Charan, your is a very good point. These cells are MSCs derived from hESCs and traditionally have been cultured onto a gelatin coating, however I tried to culture them without and they attached as well. It might be I will decide to start a full series of experiment in gelatin-free plates.

Thank you all for your valuable advices: I am now reassured that coated plates can be conserved in the fridge for a couple of days. Fantastic!

Hi Federica,

I usually coat my plates with a solution of fibronectin/gelatin, I let them completely dry (under the sterile hood) and then I store the coated plates at room temperature, even for several days/1week, and I never had problems: they work well!

Same here, we store our gelatin coated plates at room temperature and have not had any problems.

We are using 2% gelatin 225 bloom from Bovine skin. Dissolve the 2g powder by pouring it onto 100 ml cold water in a 250ml autoclavable bottle . Autoclave x 20-10 min on liquid cycle. Coat the cell culture ware adhesion surface thoroughly ,and incubate at 37˚C  x 30 minutes. Aspirate the liquid, and throughly dry at least 2h before using, and yes, the coated plates should keep for 2 weeks to 1 month 4˚C or RT.

If I trypsinize a 0.1% gelatin coated flask for about 5 -7 minutes (for splitting LMH cells), can I reuse this flask again? or maybee the trypsin manages at these few minutes to cleave the bonds between gelatin and the plastic?

Can i apply this gelatin coating for Primary RPE culture......

There is no need to leave the gelatin solution in the plate for any time at all. Put the solution in the plate and aspirate it back out immediately. As the residual solution dries the gelatin attaches to the plate.