Hello,
To those who have experience in using flow cytometry for counting bacteria (anexic cultures), I would like to know what is the detection limit.
I just had a kind of argument with a colleague about the sensitivity of FACS compared to CFU/ml counting by plating.
My opinion is that by plating we cannot obtain a "absolute" or "real" number of bacteria, and that we need to use other method such as FACS, most over when we face low concentration. This method is nevertheless suitable for comparing 2 conditions, as in both cases we are suffering the same biases, but not to obtain an "absolute" counting of bacteria contained in a solution.
In order to do so, I suggested FACS as an alternative, but I was said that FACS is not sensitive enough. My colleague claimed that culture plating is much more sensitive.
I disagree.
Please, could you let me know what do you think about it? I would appreciate some elucidation about this matter.
Thanks in advance
If you can plate out the entire solution to look for a single CFU then theoretically that would be equal sensitivity to FACS.
But if that is impractical due to sample size or other planned use for your sample there are other advantages of FACS e.g. the flow cytometer can analyse an unlimited sample volume, turnaround time to results would be faster, you can check for cross contamination by staining for your bacteria of interest, you can learn other things e.g. bacterial size to name a few.
Of course FACS is not perfect e.g. depending on the sheath fluid you use you will need to distinguish between debris and your bacteria of interest, which could be challenging for rare events (I would recommend staining with at least PI to be sure it is cells you are counting).
Each method is answering a different question. Viable counts measure exactly that. What is the concentration/ or total number of viable bacteria using the particular method you have chosen. Whereas FACS (and I agree that a DNA stain would be very useful) will give you an idea of total microbial numbers, live and dead. This would be most useful to be done with a live/dead staining method, and so could be used alongside viable counts.
- Badges
- Science method