The mice were perfused with 4% PFA. Then left in 4% PFA for about 2 weeks. The processing schedule is as follows: 70% Alcohol 15 min. 80% Alcohol 15 min. 95% Alcohol 15 min. 95% Alcohol 15 min. 100% Alcohol 15 min. 100% Alcohol 15 min. 100% Alcohol 20 min. Xylene 20 min Xylene 20 min Xylene 45 min Paraffin 30 min Paraffin 40 min. temp 58 degrees. No vac. on any of the stations.
This is the same program I use on mice spinal cords. Half the mice are under hypoxic conditions and the other half normoxic conditions. The normoxic brains also fell apart. I also did another group with the perfusion with 4% PFA then cryoprotected with 30% Sucrose. They didn't cut well. Wrinkles and air bubbles. I will try cutting at a -25 instead of -20. Also, the sections fell off as soon as I put them in water. I will admit that I have only worked with adult mouse brain and spinal cord. So this is a new area for me. Thank you again for suggestions and advice. Louisa M. Papke Research Technologist Demyelinating Diseases Research -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Geoff McAuliffe Sent: Wednesday, January 23, 2013 11:35 AM To: histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] P21 mouse brain On 1/23/2013 11:10 AM, Papke, Louisa M. wrote: > Has anyone worked with P21 mouse brain? Such as frozen or paraffin > sectioning? Right now after perfusion and paraffin embedding, the brain > literally falls apart when picking it up to embed. Any ideas or hints > available would be welcome. > > Thank you in advanced, > Louisa M Papke > > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > Perfusion with what? Fixation for how long? What is your processing schedule? Geoff -- -- ********************************************** Geoff McAuliffe, Ph.D. Neuroscience and Cell Biology Robert Wood Johnson Medical School 675 Hoes Lane, Piscataway, NJ 08854 voice: (732)-235-4583 mcaul...@umdnj.edu ********************************************** _______________________________________________ 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
