Two alternatives to the nitrogen purge for the hydrogel

I attended the CLARITY workshop, and while there, Kristin mentioned that someone was using Peanut oil on top of the hydrogel solution, or filling the tube to the top, to prevent oxygen from inhibiting the polymerization. These sounded like good alternatives to the nitrogen purge. The main problem with filling a 50 ml conical tube to the top with hydrogel, is the waste of reagent.
The first thing I tried was transferring the brains to 15 ml conical tubes and filling them all the way up to the tippy-top before closing the tube and placing in a 37 degree water bath. This worked great!! It isn't too hard to remove the mouse brains from the 15 ml conical, and it uses less hydrogel. I did have to use a metal spatula to get it lose, but it was fine. I also recently tried adding sunflower oil to the top. I didn't have peanut oil, but I did have sunflower oil on hand, it also worked great! This saves the space and cost of nitrogen tanks, vacuum pumps and desiccator. Definitely a big thumbs up.


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