Obviously the best way to avoid mycoplasma contamination is to avoid it in the first place. In our case, however, it is not possible to avoid. We are culturing viruses and tissues out of human nasal swabs, washes, and biopsies.
As you can imagine, anytime you are pulling something out of someone's nose, it is quite often full of mycoplasma. I've reviewed current examples of mycoplasma elimination, but found the section in Current Protocols in Molecular Biology to be the most helpful.
Indeed, we found that checking with a PCR based kit for common lab mycoplasma contaminants often missed detection of mycoplasma, because it did not include "clinically relevant" strains. Kits which did include these species clearly detected mycoplasma, in addition to Hoechst stain confirming their presence.
Of these mouse passage was probably the most effective, but not desirable because it might induce additional confounding factors. The kits had fair results, but 5 passages post treatment mycoplasma were again detectable via Hoechst stain (PCR detection persisted through out).
The dox and enrofloxacin may have produced better results at higher concentrations, but cell toxicity became a serious issue. I was wondering if anyone had examples of combination methods for cleaning primary cells and/or virus stocks (been grown on more standard immortalized cell lines) for a more complete treatment method.
Just as note on our hands, we regularly check our cell lines, and are in fact maintaining a mycoplasma free tissue culture system. Further, as proof of concept, we were able to clean a culture that was known to be infected with M. hominis with kits.
It doesn't seem worth the effort, but is there any evidence to suggest typing down to the species would be worth it in the eradication effort? Each donor could conceivably be providing a different contaminate.