I have cultured sf9 cell in insect XPRESS media supplemented with 10 ug/ mL of gentamicin. It looked fine for a week then it started to create a slimy looking ring inside the flask.
Hi Benjamas
I regularly get this ring when growing healthy High Five cells in Express Five media when the cell density gets a bit higher. It is just composed of insect cells stuck to the flask (dead ones, I think) and though it MAY indicate that RPM is not optimal, I have not found it to be linked to any decrease in viability or growth rate.
Hi Benjamas,
As Alison has said cultured insect cells in suspension tend to form a ring for cells round the flask at the top of the liquid level. This is nothing to worry about.
However it should not be a large ring and slimy doesn't sound quite right.
I worry that you are culturing the cells for over a week, in the same vessel. Are you splitting the cells back to around 0.5e6cells/ml when they have grown? Splitting cells every 3 to 4 days into a fresh culture vessel should keep them healthy. If not that might be why you as seeing a slimy looking ring, as dead cells build up.
Dear Alison,
Thank you so much for the message and tips!
Dear Richard, I have split cells to around 0.5 or 1e6 cells/mL very day after a few days in culture from frozen cells stock, and I have been using the same flask/bottle. I am not sure it is actual slimy but it looks like that. I have even swopped this ring and cultured in either LB broth or HI5 media, and incubate in the shaker incubator at 37C. I have not observed any growth in these 37C cultures. So I hope it is just the dead cells as you guys suggested. However, thank you so much for suggestion.
Hi Benjamas,
It sounds that's all's well.
As you are growing from a frozen stock there will be a lot of dead cells in the initial culture which could be what you have seen here. It's a good idea to change the media after a few days even if the cells don't need splitting back when you first wake up the cells.
-Richard
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