Hi, I have recently started ALI culture of Calu3 cells. Before the airlift my cells grew to a confluent monolayer with a nice TEER value, but after airlift and change to ALI maintenance media from Stemcell technologies, holes and weird cell protrusions start forming in the transwell inserts (only in some regions). This has already happened twice. I read that it can be a problem with the medium, but it is fresh, prepared according to protocol and has not met the expiry date. Has anyone encountered a similar problem? Thanks!
Dear Natalie,
I am a Product Manager at STEMCELL Technologies, and after checking in with one of our Product and Scientific Support Specialists, this is what they recommend:
“PneumaCultTM media were designed for culturing primary lung cells, and we’ve not validated them for use with cancer cell lines. Protrusions in a culture grown in PneumaCultTM-ALI Medium can be indicative of excessive proliferation; perhaps something we would expect with a cancer line. Holes may be indicative of cell death, which could be due to the differential nutrient requirements of cancer vs. normal cells.
An additional consideration is that Calu3 is an adenocarcinoma line. Adenocarcinoma is thought to be derived from the alveolar region of the lung, whilst PneumaCultTM-ALI Medium is intended for airway cells. You could therefore try culturing the cells in our PneumaCultTM Alveolar Organoid Media, with our monolayer protocol, but as these media were optimized for use with normal cells, we cannot guarantee performance with cancer cells.”
I hope this helps. If you'd like to discuss further, I invite you to please reach out to our Product and Scientific Support team at [email protected]. They will be happy to provide more recommendations once they know more about your research question.
Kind regards.
Just wanted to let everyone know that I solved this problem. I changed various things but in the end it was the Pneumacult ALI basal medium, which had gone bad. I aliquoted and froze it in the month of the expiry date and thought it was still usable, but apparently not. By using newly delivered medium, I can now grow nice ALI cultures.
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