immobilizing zebrafish

[foobytheinsane1]

Hello,

I do electrophysiology on 5 day-old zebrafish, and in order to do so I have to pin them down onto a sylgard-lined dish using sharpened tungsten pins to immobilize them. I immobilize them first with tricaine, then pin them, and then rinse the tricaine away and paralyze with alpha-bungarotoxin. The one problem which I am having is that they will often float around when I first place them onto the dish and it is then very difficult to get them pinned down as they will not stay lying flat on the bottom. Is there anything that I can do to help correct this problem?

Thanks.

[BioBM]

Pin them in low-melt agarose, then transfer them to the dish, perhaps? Pour a dish of low-melt agarose, then drop the fish onto the dish and drop more agarose on top of them, aligning them as necessary with a fine tweezer tip as it solidifies. Pin them, then cut away the agarose and remove the fish.

I used to do experiments that required immobilization in agarose, and if I sent them improperly sometimes i would free and re-set them. You have to be careful, though - it's easy to injure the fish when removing them from the agarose.