hello!
Worm papers often refer to washing of worms. Is there a standard procedure for this, ie what rpm? for how long? thanks for any advice!
We usually wash at 2000rpm (320xg) for 2 mins and the washing is repeated 3 times.
I agree, spin at 2000 RPM for 4-5 minutes. You can used M9 with 0.1% Triton-X initially to clean up any bacteria, then wash a few times with just M9 or dH20.
Hello, I’m infecting worms with different species of Candida and then mount a microplate assay with 20% BHI, 80% M9 and kanamycin. The problem presented to me is that Candida proliferates, resulting in turbidity (although I washed the worms as recommended and I allowed it to crawl on BHI agar).
I thought that perhaps the absence of cholesterol in ethanol (by an oversight) :-[ could influence. :o
If someone could guide me I would make a great favor.
Thanks.
Do washing kill the worms?
Or those worms can be further used for assays?
Hi, I’m sorry for not responding earlier.
The protocol is as described Breger et al, Briefly:
Approximately 100 worms (L4 stage) were dispensed onto a lawn of Candida and allowed to feed for 2 h. The worms were remove off the plates with M9 buffer and wash at 1200 rpm for 2 mins (the washing is repeated 3 times). The pellet of worms was allowed to crawl on unseeded BHI plates to remove yeast cells from their cuticles… but this procedure was not successful because when I pass the worms to a liquid medium in a microplate (79% M9 buffer, 20% BHI, 10 ug/ml cholesterol in ethanol, and 90 ug/ml kanamycin), Candida also grows and the medium show turbidity, and at 48 hours I can not see anything
how long should I allow the worms to crawl, so that your cuticles are well clean?
I am sorry for so poorly writing in English and thank you very much.