Oragnoid distribution

[Eric Malmhäll-Bah]

Hi Toot,

Thank you for developing and publishing the protocol. We will soon attempt to implent it in our lab. However, there is one thing that we thought we should ask about before we start: In step 30 when you add the the second layer of hydrogel containing cell aggregates to the 12-well plate it says to pipette it down the side fo the well. While doing this, do you turn the plate simultaneously or do the aggregates readily "flow" with the gel to get distributed evenly? We are bit worried that pipetting down the side of well will cause all aggregates to end up in same place? Maybe this is non-issue, but I look forwards to your answer.

Best Regards

Eric Malmhäll-Bah

University of Gothenburg

[Abdullah Khan]

Hey Eric ! The solution should be liquid enough that it will just wick across. After you've pipetted, I'd suggest tilting the plate at an angle to get the gel to evenly distribute. You can also tilt the plate back and forth to make sure you get an even distribution. If you've handled everything on ice, you will have time before anything starts to set (a few minutes at least). 

I'd also suggest doing a 'cell free' trial run just to get a feel for this yourself! 

Hope this clarifies things. In summary I don't think it has proven to be an issue - but this is a good place to check ! Let me know if it proves to be a problem, and good luck with your differentiation.

BW
Abs