I need to test their supernatant on my mammalian cell culture but it interferes with my assay. I need some media with really low Carbon source but still enough to allow the bacteria to survive (and, if possible, grow a little). I was thinking I can grow them overnight in normal media and then transfer to minimal media for some hours. However, I guess their metabolism may change as well in minimal media so not sure if it makes sense.
These are the defined medium for Lactobacillus (BL medium,Jensen and Hammer 1993), (MS13 medium, Cocaign-Bousquet et al. 1995) just try it....
You can also make up minimal MRS containing a lower percentage carbohydrate. I have used 0.5% w/v but you can go lower. The minimal MRS involves leaving out the meat extract and reducing the carbohydrate content. I have outlined the media in my paper (http://www.microbialcellfactories.com/content/10/S1/S12). You can also reduce the yeast extract concentration as well if needed.
Many thanks Dr. Prabhavathi Paulraj, I will check that out!
Michelle, thanks a millions!!! I already did some trials before see your response. I need to do some screenings of my bacteria culture supernatant on mammalian cells. The problem is that the MRS media interfer with this assay due to its high content in C source. Ideally, I would like to grow my bacteria in the assay buffer. So, what I have done is grow them overnight in normal MRS and then, grow them for 5 h in my mammalian cells bufffers assay (HBSS+HEPES) supplemented with meat extract (3%) The bacteria grow enough well (after inoculation at 1% they grow in 5 h 50% less approx. than in the full MRS media, but still they grow (OD increased)). Do you think this is good or would you suggest me still use another kind of media as that one you used?
Thanks!
Hi Cristina,
I dont really do any cell culture work myself but I had a look at the buffer media you listed and I dont think you will need to use another kind of media than the ones you already listed. You could possibly supplement your HBSS buffer with some more glucose (more than the 0.1g working solution). Again I am not sure if this is feasible for your experiment.
Many times, we saw MRS derived spent medium interfering mammalian or other co-culture cell growth. This is not only because of carbon sources, but also some metabolites. In my case, I prefer RCM (0.5% glucose) to MRS (2% glucose) for bifs, as the composition of the former is more simple. The final OD600 for bifs in RCM is around 2.5, nearly half than in MRS. I also see some researchers using half strength of MRS (or MOPS buffered) for bifs. And, if your purpose is probing some macro-molecules in the supernatant, concentrate may help.
Hi Lorenzo,
Many thanks for your reply. I cannot see the paper, sorry, could you attach it again please?
Hi Zhongke Sun ,
Thanks a lot. I will try that RCM media. Do you have any publication where you have used coculture so that I can check your protocol?
Many thanks in advance,
Cristina
Even RCM get better results in my experiments. In publications, we still report MRS as it is more widely used. http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0088260
http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0059370
I wish these papers are useful for you (see "Preparation of cell-free culture supernatant").
Hi again Zhongke,
I will check your protocols :)
Many thanks for your help,
Cristina
RCM generally is better than MRS for Bf.! MRS only really good for Bf.a.ssp.lactis
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