Organoid fusion: Day 0 to Day 5

[Beatriz Alves]

Hello,

We are experiencing issues with excessive fusion of organoids between Day 0 and Day 5 of the comBo protocol.

I have been gently shaking the 6-well plates daily to try to prevent fusion, but this has not been effective. I was wondering what volume of media you typically use when transferring organoids from the 24-well AggreWell 800 plate into a 6-well plate on Day 0.

The organoids look very good while in the AggreWell, but they begin to fuse significantly after transfer.

Thank you in advance for your help.

Best,
Beatriz

[Abdullah Khan]

Hi Beatriz - two things we've found when troubleshooting this for others: 

(1) They fuse if you have too many aggregates per well in the 6-well ULA

(2) If the incubator is a bit unstable (gas/temp fluctuations)

My suggestion would be to halve the number of aggregates within each ULA and if you have another incubator, try that. -without another incubator move the plate to a different part where the temp/gas flow is more stable. 

Thanks for tooting, 

Abs 

[Beatriz Alves]

Thanks a lot! :)