Yes, I have experienced the same thing. Many people forget about the impact temperature has on some staining reactions. If you keep your Schiffs and Periodic acid in the refrigerator it may take longer to stain than it would if the reagents were at room temp. Many old procedures are written with the reagents at room temp (even the ones requiring refrigeration). This was one of those "live and learn" situations for me. Now temp and time are the first things I look at when a stain does not work on a known control. Jan M Omaha
-----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Janet Keeping Sent: Wednesday, October 06, 2010 11:43 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] PAS staining I have for some time had a problem with Schiff's reagent and PAS staining. - I have tested each new, unopened bottle of Schiff's reagent with formaldehyde and always the color development was immediate, but purple, definately not pink.This result has been quite consistant. - PAS staining for glycogen using the method in *Histotechnology a self Instructional text,* would fail to demonstrate any glycogen in autopsy liver specimens. I teach Histology at a community college and this problem has driven me crazy for a number of years. I have tried several brands of Schiff with the same results. Recently I obtained sheep tissues which were promptly refrigerated and fixed after death, and I had hoped these tissues would demonstrate glycogen. ( My thinking was that perhaps delay before autopsy was somehow diminishing the glycogen in the specimens that I had.or that perhaps long term NBF fixation had hampered staining.) Basement membranes were stained with the Schiff reagent as expected despite the purple color in the formaldehyde test. Hotchkiss Mcmanus with the same reagent also produced beautiful staining of fungus a lovely magenta color. A search of the web made me suspiciious when I noted that Schiff added to 37-40% formaldehyde should produce a pink or red color, however, A spot check of formalin using Schiff should produce a purple color. I sent an e-mail to Brian Hewlett to ask if he could make any recommendation. Brian was not surprised by the purple color devopment in testing, He suggested that a large number of available aldehyde groups would be expected to produce this color. He suggested that I increase my periodate oxidation to 20-30 minutes and my Schiff application to 30 minutes. This worked and I am extremely grateful!. Has anyone else had an experience like this? Janet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet Sponsored by Catholic Health Initiatives and Immanuel, Alegent Health is faithful to the healing ministry of Jesus Christ, providing high quality care for the body, mind and spirit of every person. The information contained in this communication, including attachments, is confidential and private and intended only for the use of the addressees. Unauthorized use, disclosure, distribution or copying is strictly prohibited and may be unlawful. If you received this communication in error, please inform us of the erroneous delivery by return e-mail message from your computer. Additionally, although all attachments have been scanned at the source for viruses, the recipient should check any attachments for the presence of viruses before opening. Alegent Health accepts no liability for any damage caused by any virus transmitted by this e-mail. Thank you for your cooperation. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
