Mystery contamination - Tissue culture (Jun/28/2011 )

Hi, all,
There is a mystery contamination going on in my lab. We have seen small dots( about 500nm) in our COS-7 and primary macrophage cultures. Those dots are killing our cells. They move pretty fast but not Brownian motion movement. But the culture media would still be clear after overnight culture. We tried DAPI staining, it was inclusive. We tried microplasma detection kit, the results was negative for culture media and cell lysate. I have attached an image of infected COS-7 cell.
Please help.Thank you very much.
Jia

If they move and it isn't brownian motion, then it is a flagellate bacterium of some sort. Throw out your cells, medium and any other things you are using for your cultures. Get some fresh cells up.

bob1 on Wed Jun 29 01:25:20 2011 said:

Thank you, Bob. We did started everything fresh. Fresh media, fresh culture, etc.Cleaned the incubators and hood. But still get this contamination. This thing is in COS-7, and mouse macrophage, different species. DAPI showed no double DNA staining. And it is inside the cell not the culture media. Could this be some sort of virus? Thank you for you reply.

You won't see a virus with a light microscope - even the biggest ones are much too small. They could be intracellular bacteria, or they might be some form of cellular inclusion body or a mitochondrion etc.. You may need to get fresh cell stocks from another source.

Have you tested for mycoplasma?

bob1 on Wed Jun 29 09:51:41 2011 said:

Thank you Bob.
The image was taken using 60X 1.49 oil lens. The resolution is about 130nm. So, the size of the particle is about 300-500nm. Yes, we tested mycoplasma. It was negative. We are getting fresh cell stock and culture media from another lab. I think it might be the FBS we used somehow got contaminated.