When initiating a primary culture, what is the least amount of time antibiotics/antimycotics should be used? Would just a few hours be sufficient? In addition to issues affecting viability, etc., I also want to avoid antibiotic/antimycotic resistance. What length of antibiotic/antimycotic administration would be considered "too long", in that the potential for the development of resistance might be invoked? As far as I could find, there doesn't seem to be an answer to the latter question. Agricultural studies have found antimycotic resistance to have emerged after a growing season, however this doesn't take into account when during the growing season resistance may have arisen. If anyone has any insight, I'd appreciate it. Thank you.
DEAR KESSEL
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THIS BOOK MIGHT GIVE YOU SOME USEFUL INFORMATION.
Studies on Experimental Toxicology and Pharmacology
edited by Stephen M. Roberts, James P. Kehrer, Lars-Oliver Klotz
Interesting, basic question. In that cell lines are routinely passaged for tens or even hundreds of times with antibiotics/antimycotics in the media, I'm not sure that there is an upper limit for the duration of use. For a primary cell culture, I would think that use for the duration of the culture would be fine for most cell types. Certainly, I've never had issues with myoblasts, neurons, or glial cells, though as with most researchers, I've also not looked for issues that may be there aside from the appearance of obvious contamination. If the concern is the appearance of antibiotic-resistant bacteria (or antimycotic-resistant fungi) appearing in the culture, I think this is a minor concern. When I first learned to culture primary neurons, I never used antibiotics/antimycotics and I never had contamination issues; I just learned good sterile technique practices. Culture performed using proper sterile technique should address most contamination issues. Since antibiotics often don't kill, and only prevent bacterial proliferation, removal of them from the culture may make any bacteria become readily apparent if you did not use good sterile technique to begin with.
Dr. Singh and Dr. Young,
Thank you for your replies. It's reassuring, at least, that there probably won't be any issues if treatment is used for the duration of a culture.
When I received primary tumour tissue, I collected in transport media (RPMI + 2% Pen/Strep/Amphotericin B) and washed the tissue 3x10mins on a roller, changing to fresh transport media each time. I then disaggregated and plated out in antibiotic/antimyotic free media from then on and generally didn't get any contamination. I was collecting from the oral cavity as well which has a high bacterial/fungal environment
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