The volume of a mouse is about 25ml, of which perhaps 2ml is blood. 5ml of saline (or PBS) is enough to wash it all out. Some people precede this with a small bolus (about 0.2ml) of 1% aqueous sodium nitrite to dilate the blood vessels. After washing out the blood, perfuse with about 10 ml of fixative, over the course of 5 to 10 minutes. It's important that the pressure isn't too high, to avoid rupturing capillaries and small veins, and waterlogging (wrecking) the tissues. If perfusing from a syringe, use the thinnest available needle to attenuate the pressure; 10 minutes feels like a lot longer when you're pushing the plunger very slowly and keeping the sharp end of the needle motionless! John Kiernan Anatomy, UWO London, Canada = = = ----- Original Message ----- From: Li Zhang <dancingw...@yahoo.com> Date: Wednesday, March 17, 2010 14:59 Subject: [Histonet] question about mouse perfusion To: histonet@lists.utsouthwestern.edu
> Dear all, > > I have just started to learn how to do perfusion in mice and > this site has helped me a lot. > > We don't have the pump or Perfusion one in the lab, and probably > won't be able to get one in the near future, so I'm using the > hand injection method. I'm using a 60 ml syringe with 21G > needle. I am doing perfusion in order to do immunofluorescence > on the brain. > > My question is: can anyone give me a rough idea of how fast I > should inject ( like ml/min). I think I've tried like 30 ml in 3 > min, and I suspect that it's too fast because I do observe > tissue swelling sometimes. > > And another question, which of the following is better for > perfsusion: 1) PBS and then 4% PFA 2) Saline and then 4% > PFA 3) sucrose--> saline--> PFA? > > Thank you very much for your help! > > > > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
