Planning to use Ice cold PBS protocol (Mahé, Maxime M et al. “Establishment of Gastrointestinal Epithelial Organoids.” Current protocols in mouse biology 3 (2013): 217–240. PMC. Web. 19 June 2015.), but i saw some comments that we can use dispase solution to remove matrigel instead of PBS or Recovery solution.
need some help from you guys!
I have been working for a while in spheroid culture and analysis. To my experience, TrypLE express works better than dispase. If you plan to use your culture for further experiments, do not use ice cold PBS protocol which decreases the survival of cells.
Hi Parthasarathy
Thank you for tip. It is really good to know before i use ice cold PBS. I'll try with TrypLE express.
If you are just splitting the cells then I have found cold PBS fine - use the P1000 pipette tip to break it up mechanically a little. Then centrifuging at increasing speeds should isolate the organoids etcs. If you are wanting single cells then 5 mins in trypsin (37 degrees C) should do it. Hope this helps
