Hi everyone,
Maybe this topic was already adressed but I couldn’t find a post about it.
I’m desperately looking for a protocol to efficiently immunostain a small quantity of worms. They are very sensitive so freeze-cracking is not an option, unless you know how to avoid them bursting each time.
I would like to use a finney-ruvkun type of protocol, but on a few (10-50) worms it’s probably useless, I’ll have no worm left at the end…
Any idea?
Freeze-cracking with poly-L-lysine-coated slides is usually fine for a small number of worms. If you have too few of your specimens and are worried about ‘oversquishing’ them, add some animals of a morphological mutant that you can easily distinguish from your worms, like Dpy animals.
Hi, and thanks for your answer.
So you mean, adding a few dumpy for instance will “cushion” the slide and help the coverslip from squishing?
I used superfrost charged slides, worms sticked fine but got squished, so I’ll try your method for sure!
What are you staining for? I assume you are dissecting them to release the gonads, so you are trying to look at the intact worm? I think the previous persons suggestion of using some dpys to help cushion should help you.
Mix your precious worms with an excess of GFP expressing ones (make sure that the GFP survives fixation) and proceed with the Finney-Ruvkun protocol. I used that with great success.
Thanks for the tips !
I tried to extrude the gonads by dissection, they never come out and/or explode.
Freeze-cracking make them burst, but I might try cushioning with Dumpy worms.
I think doing the finney-ruvkun with an excess of GFP+ worms might help, I’ll try all that thanks!