Hi everyone. As part of an exposure, I need to feed C. elegans cultures deactivated E. coli OP50. We've had little success in deactivating the OP50 using freeze-thaw cycles and heat inactivation. Also, exposure in a UV box did not produce great results. The problem is that I either need to deactivate a washed OP50 pellet, or a suspension with an optical density of 3 (at 595 nm). So a very high concentration! Does anyone perhaps have any suggestions? Maybe a UV inactivation protocol (with details about exposure time, light intensity, what type of containers, etc.)? Or maybe autoclave at lower temp and pressure? Lysis should of course be avoided. Your help will be much appreciated! Thanks.
I just have suggestion, I do not have any experience, you can yous bacteriostatic agent like Chloramphenicol .
Hello, chloramphenicol and UV can not fully deactivate E.coli even in concentration mentioned. Also adding some chemical or antimicrobial agent will not allow to use E. coli as feed in future. So the better way that you mentioned heat treatment if you want to deactivate culture without total lysis.
Thanks for the replies. Alirenza, we have been using Pen-Strep, but I would like to avoid the use to high concentrations of antibiotics. Karen, do you perhaps have a protocol? Heat treatment (at 65 degrees celcius) did not deliver good results in the past.
70*C 40 min will be enough for heat treatment. Put suspension in 70*C water bath for 40min mix periodically.
I know from my experience in isolation of spore forming microorganisms from soil. After heating at mentioned conditioned no any vegetative living form observed.
Karen A. Darbinyan
that temp and duration you mentioned cause of lysis bacteria, is not?
Most part of microorganisms should avoid lysis if your suspension pH is in a range from 6 to 7.5.
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