Our packaging cell line was infected with mycoplasma, and our virus-containing media also contains mycoplasma.
It is recommended to discard it, but is there any other way?
We've thought to centrifuge the media and to remove the supernatant, and to add Mq-water to lyse the mycoplasma.
They should "explode", but after that, how can we check if mycoplasma DNA remains via PCR?
If your packaging cell line is mycoplasma contaminated your lentivirus titer will be very bad anyhow. The best you can do is discard these virus and start again with a line that is mycoplasma free. You might try to make it mycoplasma-free with Ciprofluoxacin on any of the antibiotics they recommend above, but to make sure they are mycoplasma free you should subclone them after the treatment and get one clone that is mycoplasma-free. Otherwise ask your colleagues for the packaging cell line mycoplasma free, but check it anyhow. One of the characteristics of mycoplasma is that the transfeccion efficiency of the cells decreases at least one log. This is the reason why I mention that the titer of virus will be very low.
Hi Daniel,
If you can not replace virus producer cell line, you may try to treat cells with Plasmocin or Plasmocure (Invivogen).
Invivogen have pretty good kit to test mycoplasma contamination as well.
Good luck!
Lena
I had better results with BM cyclin from Roche than Plasmocin. But maintaining in Plasmocin will help during clearing other cell lines in the same incubator. Other precautionery measure is to use Aquagaurd from Promocell to keep your culture clean. Good luck!
There are 0.1 um filters rated for mycoplasma removal. You could fish them out with one of those, and your virus would stay in the filtrate.
If your packaging cell line is mycoplasma contaminated your lentivirus titer will be very bad anyhow. The best you can do is discard these virus and start again with a line that is mycoplasma free. You might try to make it mycoplasma-free with Ciprofluoxacin on any of the antibiotics they recommend above, but to make sure they are mycoplasma free you should subclone them after the treatment and get one clone that is mycoplasma-free. Otherwise ask your colleagues for the packaging cell line mycoplasma free, but check it anyhow. One of the characteristics of mycoplasma is that the transfeccion efficiency of the cells decreases at least one log. This is the reason why I mention that the titer of virus will be very low.
I would second Jose's answer. We had this problem one time, and virus production is very poor in contaminated cells. If you can re-make the virus, that would be ideal...
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