We have been growing cells in 3D organoids (1:1 matrigel/growth media mixture) in a transwell insert. However, when we try to fix the cells with 3.7% PFA, the matrigel becomes a little loose and we are afraid that we may loose the 3D structures of the colonies. It also makes getting the colonies out of the insert a challenging.
We would ideally like to embed these cells in OCT media for subsequent sectioning and staining.
Hi Jaymin,
we normally fix hydrogel based 3D organoids in 4% PFA, for OCT embedding we perform sucrose aascent from 10 to 30% and then snap freeze in OCT.
Good luck
Hello
Yes , use 4% PFA, for OCT embedding and perform sucrose ascent from 10 to 30% and then snap freeze in OCT , Its a good method .
Good Luck
Check this paper. Hope it helps.
https://www.ncbi.nlm.nih.gov/pubmed/30548444
Generation of Pancreatic Ductal Organoids and Whole-Mount Immunostaining of Intact Organoids.