You can't go from 70% alcohol into paraffin without passing through 100% alcohol and then a clearing agent (liquid miscible with 100% alcohol and with melted wax). Xylene is a commonly used clearing agent. Your times in 50% and 70% alcohol are much longer than necessary. Even for a whale's hypothalamus a few hours in each solvent step should be adequate. In Belgium you may be able to obtain a great classic of histotechnology: Gabe, M (1968) Techniques histologiques. Masson et Cie, Paris. 1113 pages! This explains tissue processing very thoroughly. The author was an academic zoologist who did all his own lab work and made sure he knew what he was about. Get a copy if you can. The English translation of Manfred Gabe's book was posthumously published in 1976. I bought one then and learned a lot from it. Like many histotechnology classics, this book is now a prize possession, almost unobtainable on web sites for second-hand books.
'nuff sed. John Kiernan UWO = = = > -----Original Message----- > From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet- > boun...@lists.utsouthwestern.edu] On Behalf Of An Eerdekens > Sent: Friday, January 28, 2011 4:18 AM > To: histonet@lists.utsouthwestern.edu > Subject: [Histonet] paraffin sections: white with airbubbles > > > Dear collegues, > > I am experiencing following problem. > > I have embedded hypothalamus tissue in paraffin using the > following procedure: -fixation in 4% paraformaldehyde for 48 > hours, fixation of the tissue in 50% alcohol, next day in 70% > alcohol,next day paraffin embedding. During the paraffin > embedding there was a short circuit and the machine did not work > for any hours, so there was a delay in the process. > > Now I am making slices of 5 micrometer, using the Microm HM 360. > > The tissue is very white (looks like I am making much thicker > sections) on the slices with airbells inside. I don't have an > explanation for this and many samples are showing the same features. > > Does someone know what might be the reason? > > Thanks for the help. > > Regards, > > An Eerdekens > Laboratory of Intensive Care Medicine > Catholic University Leuven, Belgium > > 003216330518 > _______________________________________________ > 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 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
