I'm currently a research intern culturing iPSCs and have been encountering a recurring issue with colony detachments and unusual debris in the media. Despite my efforts, I haven't been able to identify the root cause and would greatly appreciate any insights or similar experiences from this community.
Here's a brief overview:
- My PI instructed me to discard the initial affected plate, suspecting contamination, but the mycoplasma tests returned negative.
- I received fresh cells from a colleague, yet the phenomenon persists, even after multiple passages.
- The detachment always initiates at the bottom region of each well and gradually spreads. In one well, nearly a third of the colonies detached.
- I initially suspected issues with plate coating (Geltrex), but other lab members using the same cell line and protocol do not experience this issue.
- I've attempted to aspirate media carefully from the top side of the wells during media exchanges, but the detachment consistently originates from the bottom region.
Has anyone encountered a similar phenomenon or could suggest potential troubleshooting strategies?
Thanks in advance for your help!
Hello,
I am currently facing the same exact problem, with colonies detaching from the bottom area and increasing cell death. I am currently looking for an answer myself. Did you manage to find a solution?
I have the same problem. How do you passage them? Do they detach when you change the media?
Dear all,
I am not sure if this helps, but I would try this:
1. ask your lab colleagues to guide you with the iPSC culture. If they do not have that problem, maybe it is just because of the handling. iPSCs are quite sensitive.
2. You did not tell us when, after the passage, you made the pictures.
3. Which type of passage do you use? Single cell split or colony split?
As a quick response, I would say they are stressed. I would try to increase the cell density after passage. Then I would check the media and coating if they are okay. The contamination is still in game, could be also bacterial.
I hope you will resolve your problem quickly.
Mr Eunsu Lee have you found the reason of these bubbles? I'd like to know it too
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