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There could be two issues. If by “depleting” you mean that the volume of media is going down, then this could be an issue of evaporation. If by “depleting” you are speaking of nutrient depletion, than this could have a few different causes. If this is the case you should check the CO2 and water levels in your incubator to make sure that these are okay. If you exhaust all these possibilities and suspect contamination some quick tests would be:
- Check by PCR and microscopy to make sure that your cells are consistent in their morphology and genetic profile.
- Bacterial: Visual turbidity and pH of the media.
- Endotoxin: Limulus amoebocyte lysate (LAL) assay;
- Mycoplasma: Hoechst stain