Hi all,
I am trying to make an IPSC-derived bone marrow organoid.
I did flow cytometry on them after they are made at Day 10 maturation. The bone marrow organoid has erythroid cells, endothelial cells but are lacking in hematopoietic cells with no CD45+ signal. I believe that they did go into mesoderm commitment because at Day7 when I tested them via flow cytometry, they were CD34+ and CD31+.
I am troubleshooting why this is. There are a few problems I could address:
-IPSC health is poor. I will try to add Revitacell during Embryoid Body generation.
-Some of the cytokines I used to create them may have degraded. I accidentally vortexed them for 30secs when reconstituting in sterile water. I have an older batch of cytokines that I know were not vortexed. I will use these to compare with the new cytokines.
Anyone with experience in this? thanks!