I am looking at alternative options for quantifying bacterial viability in addition to colony plate counting. I have a mixed species bacterial culture that may also contain epithelial cells. I have tried ATP as a metabolic measure. The challenge I have is correctly quantifying bacteria ATP vs epithelial. Is there a way to filter or eliminate everything but the bacteria? How can I make sure I read only the bacterial ATP?
The epithelial cells should not be viable in LB for long term, I image they should die once they are cultured over night in LB and only bacteria will be left. If you are not sure you can also put the entire solution in media in a TC flask. The cells should adhere to the plate within a couple of hours but the bacteria should not. At that point you can take off the bacteria only suspension and do MTT on that. Of course a few cells could slip through but likely most everything will be bacteria.
You can try to kill eucariotic cells by incubating your samples with reasonable amount of Cycloheximide and then you can use AlamarBlue assay for evaluation of viable bacteria in your samples
After cycloheximide treatment you can check the bacterial cell viability by ITZ assay.
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