I'm cultivating suspension mammalian cells in a bioreactor, I'ld like to know what all the methods to detect/know whether it is contaminated or not.
Few points from my side are :
1) DO (Disolved O2) level rapidly drops.
2) pH rapidly drops.
3) media color changes.
4) Huge turbidity.
Apart from above points are there any other ways/methods through which we can detect/spot/identify contamination in mammalain cell cultivating Bioreactors. I was just curious to know the others ways also.
Hi Vikram Kumar
Here are some points from my side:
You can also check for:
1) High levels of ammonia.
2) Analysis of the exhaust gas can be equally effective for detecting the presence of undesired respiratory activity.
3) Metabolic footprint analysis (exometabolomics). The metabolic footprint is the profile of extracellular metabolites resulting from nutrient uptake, extracellular biochemical reactions, and the secretion of metabolic products performed by a population of cells in the environment in which they grow.
4) For cell cultures contaminated with mycoplasma or viruses, there may be no visual changes to look for, even if using a microscope. Poor cell growth and overall performance of the culture may be the only clues that a problem exists.
Best.
Hi! As it was mentioned before, there are some typical signs of contamination: low pH, high oxygen consumption (this phase can be 3-6 hours long, before your DO-controller reach it's limit and DO will drop down), high lactate(!) and ammonia in some cases.
Both bacterial and fungal contamination can be easily spotted via simple microscopy of a samples.
With viral contamination it is much harder to clarify an exact reason for your cells to die or stop growing. The best results we obtained by simple DAPI staining of sample.
And you should never hesitate to ask your colleagues about previous history of your room, where bioreactor is placed - it could help you to get an idea, of what virus you should test for. Simple, but very stupid fail from my own experience - I worked with HEK293 cells in the same room, where people before me worked with replication-defective adenoviruses.
Dmitry Lifanov, Does DAPI staining will inform about Viral Contamination ?
Vikram Kumar sorry for late answer, I was a little bit busy) In case of adenoviral contamination infected cells had quite bright cytoplasm staining, not only a nucleus.
Still viral contamination has been a big questionable part in bioreactor contamination. I would suggest to go with a small scale purification (if the batch is suspected to be infected with viruses which you particularly don't know). Include "cellufine cellulose" resin to purify the virus followed by a HPLC analysis to confirm whether there is a match in the viral detection parameters.
Someone else said low pH but also high pH if it's a fungal contaminant. So observing the color of your media is key.
Things only get cloudy if the contaminant is far along.
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