Hi Cherie, Since the top section is consistently better, then the possibilities lie in what happens after they are on the slide. As Toni recommends slides that are heated or deparaffinized vertically may have left over paraffin on the lower regions.Maybe you could also try depar with them up on their long edges. Staining slides vertically might also give this appearance if reagent times and rinsing are too short. Staining slides flat would point to slides not level or reagents not evenly dispersed on the slide. I would not rule out thick-thin on ribbons but that is probably not the case since the good section is always at the top. Good luck, Tom T
-----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Rathborne, Toni Sent: Wednesday, December 04, 2013 10:17 AM To: 'Chapman, Cherie J.'; histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] (no subject) Could it be the heating/deparaffinization process? If the upper sections are staining more evenly, then maybe they are free from residual paraffin. Try extending the time in the ovens and/or xylene. -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Chapman, Cherie J. Sent: Wednesday, December 04, 2013 1:12 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] (no subject) Hello all, I am looking for suggestions on issues with our H&E stain. I supervised a Veterinary Diagnostic lab for over 27 years and produced top quality sections, H&E's, special stains and IHC on a variety of different species in our lab. I am currently working in a Dermatopathology Lab and I am finding inconsistent staining with our H&E's. Working with just skin is a challenge all on its own. We have made changes to our staining protocol and just not happy with the end product. What we are observing is inconsistent staining on levels on the same slide. The top section seems to stain more evenly than the middle and bottom sections. I can actually see three different shades of color. The specimens are ribboned sections so I know it is not from thick and thin sections. We have looked at our processing times, microwave vs. oven times, staining reagents, different brands of hematoxylin and eosin, adjustments on staining times, tap water compared to distilled water. Our main processor is the Thermo Scientific STP-420 and our back up is the Sakura VIP V processor. I have been working with Thermo technical support thinking it might be a processing issue. We have a Leica ST5020 Multstainer/CV5030 Robotic Cover slipper we have made several changes that the technical teams has suggested to the reagents and staining time. It's still not the quality that we are looking for. I have had culligan techs out several times to see if it could be something with the water. We can run 100 slides the same day, same reagents and protocol and the H&E color is so inconsistent. I would appreciate any suggestions in this matter. Cherie Chapman, BS, HT, HTL (ASCP) Associate Director of Dermatopathology Laboratory University of Missouri Department of Dermatology University Physicians Medical Building Phone: (573) 884-0123 Fax: (573) 884-0834 _______________________________________________ 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
