drop fixation for CLARITY

jburn

Has anyone drop fixed brains for CLARITY instead of perfusing? I have neonatal brains that I would prefer to drop fix instead of perfuse but am worried it won't work? If you've done this, does it work and how long did you drop fix them (and did you do acrylamide and PFA or just PFA initially?). Our lab uses the PACT/PARS protocol.

Adam

I haven't done it on neonatal brains, but on adult, it works fine. However without perfusion, you'll likely end up with unwanted fluorescence in the blood vessels. On adults, I've used PFA for 24 hours at 4 deg, followed by hydrogel for 7 days, or alternatively just hydrogel for 4 days. Both seem to work fine, so you could just reduce the time for neonatal.