Hello,
Are there some tips for orienting the worms on their front or back so that I can see left and right axons or neurons? Worms mostly are positioned laterally on the agar. Is there any any way I can roll them over?
Best,
Faten
Hi Faten,
I had the same problem looking at muscle arms in worms. Ended up glueing them on an agar pad with cyanoacrylate glue. Simply dabbed some on the nose (using pick) first to keep the worm ‘still’ (ish) and used more glue to twist the worm ventral side up (by looking at vulva). Was rather tricky though, and on the standard microscope the glue was autofluorescent when looking at GFP (not good!) although the confocal seemed to filter it out well. I think there are other glues available that may not be auto fluorescent e.g. http://www.wormbook.org/wbg/articles/volume-18-number-1/worms-get-a-brand-new-glue/
I spoke to some others who have looked at ventral nerve cords and they say they simply put lots of worms (e.g. 60) onto the pad and took images of the 5 or so worms that ended up facing the right direction, this may be easier for you if numbers aren’t a worry.
I’ll be interested to see if anybody has any other methods to flip worms on their front or back. I seem to recall something about a microfluidic chip where one could immobilise worms and then turn them around… Not sure if it was just a dream as a (very) quick google search yielded no obvious results…
Good luck,
Ben
It is possible to “roll” a worm immobilized on an agar pad by drawing the cover slip across it in the desired direction of rotation. It’s frustrating, though.
You can also use Rol (roller) worms for some applications: because the entire animal is twisted, it cannot lie entirely on its side: if it’s on its side at on point on the anterior-posterior axis, it won’t be on its side some distance anterior or posterior of that point. Obviously, this isn’t a good solution if you need to look along the ventral or dorsal surface for any great distance along the animal’s length.
Yes people also told me about transferring a lot and using the few that are rolled over; it will be my last resort as I want a lot of worms. Oh gluing, I didn’t think of that. Thank you for sharing the link about glues, that is another idea that I will try. As for the coverslip rolling, it is indeed frustrating, yes. But I think the rol- mutant is what I will also try as I don’t think I need to look at long lengths.
I tried to just slow the worms down but not immobilize them, the worms head would move a bit, and I could see from the top. One problem is in taking a picture, as there is a lag between scope and camera.
I will be trying these suggestions! Thanks you guys!
Best Regards,
Faten
I have had wonderful luck paralyzing worms using a few microliters of 10 mM muscimol on the pad. It takes a little longer than levamisole, but the worms really stretch out and look like little logs. They also roll a little easier when you put the coverslip, allowing some of them to be ventral side up. They also aren’t as twitchy under the confocal.
That is very interesting! I will check if we have it in our lab, and if not very expensive, maybe ask to order it. Thank you for sharing.