Preventing contamination in longevity experiments without FUDR

Hello,

We are trying to run some longevities without FUDR and we keep getting various forms of contamination on our plates. Any tips other than just being as careful as possible with keeping reagents sterile? Our worms are bleach synchronized for each trial.

Thanks,
Keith

What kinds of contamination? Getting fungal contamination is inevitable and the experiment can go on if you ‘clean’ up your experimental worms by transferring to new plates 2-3x (wait an hour or two between each transfer) as soon as you notice the fungus. The fungal contamination shouldn’t affect your assays if you clean them up quickly.

I usually put a antibiotic and antimycotic mixture into the NGM medium (penn, strep and amphotericin B). Add this after cooling the NGM shortly before pouring the plates.
Worms do fine in on the plates and we have very little problem with contaminations.

Usually I just make three plates for everything and end up censoring a few contaminated plates out of each experiment. Since the contamination tends to show up later on, after a lot of worms have died, that might mean censoring 5-10 out of 100 worms.

You can also watch for signs of contamination and transfer the worms to a fresh plate before the contamination spreads with a bit of luck. Little fungal colonies haven’t made spores that will contaminate the new plate. Sometimes you can spot an isolated bacteria or yeast colony and transfer the worms before they start spreading the contamination. However, I don’t like transferring just a small subset of worms, since you may be causing picking injury to just one of the experimental groups.

Other than that I just do my best to throw out moldy plates at the first sign of contamination, and don’t sit next to labmates when they’re digging through their old moldy boxes. I wonder if some sort of air filter might reduce the airborne particulates that cause a lot of contamination…

Thank you very much for the suggestions. We can get both fungal and bacterial contamination, so all your comments should help.

Marlon,

Can you tell me the concentrations of penn, strep, and amphotercin that you use?

Thanks,
Keith

When I am lazy I usually use a 100x concentrated mixture from Gibco
Antibiotic-Antimycotic (100X), liquid
http://products.invitrogen.com/ivgn/product/15240062

Good luck,
Marlon