Sample collection from culture of facultative anaerobic bacteria growing anaerob - (Aug/30/2008 )
I am working on Klebsiella pneumoniae, a facultative anaerobe. I have to grow it anaerobically (100ml culture), add copper solution (400ul) at mid log phase and then take samples (5 ml) at regular intervals of time.
How can I do so without disturbing the anaerobic environment? Please guide me in detail.
I do not have anaerobic chamber.
Thanks
You can use serum bottles (google it). They should come with stoppers that can be sealed airtight. You can use a syringe with a needle to take samples. The rubber will keep the bottle sealed. We use that for anaerobic tuberculosis cultures.
Thanks a lot UGA80.
You have told me a very simple solution. But if I draw out sample once, there will be vaccum inside the bottle. Will it not cause any problem. I have to take samples 5 or 6 times.
Probably depends of the size of your samples. But I don't think that you will create much of a vacuum. But as far as I know, that is the only solution I can think of.
Changing the headspace as a function of repeated sampling would be an uncontrolled variable. Prob. not important but unnecessary and not good science. Maybe you could technically address the potential variability it would introduce. How have you established anaerobic conditions and what are you trying to measure with this sample event?
suggest you could culture identically in multiple vessels and "sacrifice" one for each of the sampling events.
suggest you could culture identically in multiple vessels and "sacrifice" one for each of the sampling events.
I have not yet grown it anaerobically. I am planning to do so. I will fill the bottle completely with the medium. thus bacteria will have consumed any oxygen present in the medium till log phase. Then I shall add copper solution and take samples. These samples will be used for copper estimation per cell dry weight throuh atomic absorption spectrophotometry. I have done it under aerobic conditions. More over these samples will also be used for determinig the effect of copper on expreesion level of some related genes under aerobic and an aerobic conditions.