Davidsons fixative is popular for eyes. This has been discussed at great length on Histonet, so search the Archives. I believe it works for IHC. Gluteraldehyde can crosslink antigens too strongly if IHC is needed. Retina commonly detaches, at least with larger eyes we have worked with, when the eye section is flattened on a water bath. The water bath temperature used was a few degrees lower than normally used for other tissues. You should drain the slides well in upright position if using Plus charge. Lay flat on slide warmer, to dry at 37C to 40C overnight or longer as we do for decalcified bone sections.
There are other ways to flatten eyes by floating an eye section on 5% alcohol water bath, then picking up on a slide, and slowly lowering the section (keep top part of paraffin of section attached to slide, and lowering this into a warm waterbath. The surface tension is reduced by the alcohol, allowing the eye to very slowly flatten, then pick up, drain well then dry flat. This takes a bit of practice but also works for decalcified bone when the softer cartilage likes to wrinkle versus the harder bone. Always pays to slow down speed of water rinsing, if the water flows too hard, during staining. Gayle Callis HTL/HT/MT(ASCP) -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Roger Moretz Sent: Monday, November 23, 2009 2:15 PM To: Durden, Kelley; histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] Mouse eyes Altho' retired for 2 years, we had a protocol that retained all parts of the eye quite well. Fixation was in 3% glutaraldehyde (diluted in H2O), 4 deg.C for overnight. Don't extend the fixation as this will cause the tissue to be too hard. Wash for about 1hr in running tap water and process as usual for mouse tissue (if it works for mouse liver, it will work for the eyes). Roger Moretz, Ph.D. (ret.) ----- Original Message ---- From: "Durden, Kelley" <kelleydur...@pathology.ufl.edu> To: "histonet@lists.utsouthwestern.edu" <histonet@lists.utsouthwestern.edu> Sent: Mon, November 23, 2009 10:44:21 AM Subject: [Histonet] Mouse eyes My question concerns mouse eyes. Can anyone send me their fixation suggestions? We are receiving mouse eyes that end up with a "wavy" look instead of a nicely defined "C" and have no explanation for this occurrence. Can anyone send me their mouse eye processing schedule? We have a protocol that has proved fast and true but would welcome other suggestions. Can you give me a good idea for making sure sections stick to slides well? Gold Plus? What else? Has anyone experienced a retinal detachment after or upon staining? Retina attached after sectioning - detached after staining - just routine H&E. Should I heat for an extended period of time? Kelley Durden, HT ASCP University of Florida Molecular Pathology Core _______________________________________________ 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 __________ Information from ESET Smart Security, version of virus signature database 4630 (20091123) __________ The message was checked by ESET Smart Security. http://www.eset.com __________ Information from ESET Smart Security, version of virus signature database 4630 (20091123) __________ The message was checked by ESET Smart Security. http://www.eset.com __________ Information from ESET Smart Security, version of virus signature database 4630 (20091123) __________ The message was checked by ESET Smart Security. http://www.eset.com _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet