Thank you all very much for your suggestions. I'm going to play around with a progressive H&E when I return from vacation next month. I do have safranin on hand but will need to order some phloxine to experiment with. I will probably need to order some additional supplies to make the hematoxylin. I'm not sure yet if I'll start with Mayer's or Gill's or maybe even Erlich's.
I should note that cutting at 2 isn't required, but it is desired once they saw that I can. And since I can, I aim to please! :-) In addition, I'm looking forward to trying the oven dry method before coverslipping. A rapid dehydration isn't really possible since I'm working with a xylene substitute (Pro-Par) and have been battling eosin carryover but that is a whole different animal for another thread. Thanks again! I'll report back in a month or 2. On Wed, Jun 29, 2016 at 10:27 AM, Rene J Buesa <rjbu...@yahoo.com> wrote: > > Angela: > "Pale" results are the trade-off for great quality very thin "2 µm" > sections but you can always improve intensity somewhat . > 1- your "regressive" stain, if it is "modern Harris" has the inherent > problem of lacking mercury chloride and it is little you can do about. > Perhaps if you use "progressive Mayer" you will get better results. You > will not have to differentiate (with the intrinsic "danger" of leaving the > section too pale) and if used fresh Mayer's can be a good approach. > 2- as to the counterstain perhaps you should add safranine to the eosin > (20% safranine + 80% eosin) and will get a darker red. > 3- try to dehydrate as quickly as possible or even better, wash the > sections in distilled water and place them in an oven at 60ºC for 10 > minutes and coverslip as usual. You will eliminate any "color wash" due to > the alcohols. > If you've not enough "trust" on dry/oven dehydration, try with some > sections as a test. You will like the method. > René > > > On Tuesday, June 28, 2016 5:53 PM, Angela Lamberth via Histonet < > histonet@lists.utsouthwestern.edu> wrote: > > > When I cut at 2μm my H&Es and special stains look pale. How can I get my > stains to pop or am I stuck with pale looking stains when sectioning that > thin? > > I run manual specials and a manual regressive H&E. For H&E I've tried > increasing my time in hematoxylin (beyond the manufacturer recommendation), > diluting my acid alcohol differentiation, and increased time in eosin but > the slides still lack the vibrancy that many of the postdocs desire. > > I use Shandon instant hematoxylin and alcoholic eosin by Thermo. Everything > else I prepare in house from scratch. Any recommendations? > _______________________________________________ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > > > -- Angela Lamberth Histology Technician II Histology Core Lab La Jolla Institute for Allergy & Immunology 9420 Athena Circle La Jolla, CA 92037 _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet