In my human liver cell lines, at 10x I see many black dots floating in the media. At 20x, I see them moving around (but just within a small area). They are basically just hanging out outside of the cells. A few things are odd about this:
1.) The cells are being kept in 1% penicillin (from 5000 IU/mL stock) and streptomycin (from 5000 ug/mL stock). I have since increased the antibiotic to 1.2% with little effect.
2.) If they are bacteria, they never reach log phase of growth. I have kept a flask for over four days with them. The media does not become turbid, and the number of dots remains constant.
3.) I have split off
Hi,
it is very hard to distinguish non-motile bacteria from e.g. protein aggregates or vesicles shed by the cells, as both move similarly due to brownian motion. Can you provide an image or even a video clip of your culture?
As you suspected, if you have a resistant contaminant, it is very likely that they do not reach log phase, as most resistances increase tolerances towards antibiotics, but do not completely abolish their effects, possibly leading to non-obvious lingering contaminations.
To check for that, simply withdraw antibiotics and observe the culture for a few days, if you have bacteria in there, they will overgrow your culture quite quickly, being easily recognizable already by medium turbidity only. If so, throw out your cells and start with fresh stocks.
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