Frances is using the classic formula for Schiff's Reagent. I have done the activated charcoal step for a few hours and overnight if we started too late in the day. Both times should work very well. Thank goodness we can now buy from good vendors that ship good Schiff's. I have been bright pink more than once, and it is not my favorite colour. We use the same vendor she uses by the way.
Pamela A Marcum University of Pennsylvania School of Veterinary Medicine Comparative Orthopedic Laboratory (CORL) 382 W Street Rd Kennett Square PA 19438 610-925-6278 -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Swain, Frances L Sent: Tuesday, February 17, 2009 10:32 AM To: pbru...@siue.edu; histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Problem with Schiff's reagent Sorry Paul I replied to Renee in stead of you. I purchase my Schiff's from PolyScientific R & D. When I made my Schiff's Used essentially the same recipe that you do. I made mine in a flask stored it in the dark for 24 hours. I then added activated charcoal to excess and let it sit in the dark for at least 2-3 hours. I then filtered it through filter paper that to which charcoal was added to the filter paper before I pour the Schiff's in it to clear. It will usually clear up if it has a slightly yellow tinge you can put it in the refrigerator in a tightly capped bottle overnight and it should become clear. It is important to store your Schiff's in the refrigerator when you are not usuing it and use a brown bottle. You can test your Schiff's to see if it is viable by dropping a drop of your Schiff's in 50 ml of 37-40% Formaldehyde. If it turns purplish pink it is still good if it turns any other color. Do not use it. Frances L. Swain HT(ASCP) A. A. S. Special Procedures Technician Department of Orthopaedic Surgery Center for Orthopaedic Research Barton Research Building 2R28 4301 West Markham Street Little Rock AR 72205 (501) 686-8739 PHONE (501) 686-8987 FAX swainfranc...@uams.edu email -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of pbru...@siue.edu Sent: Saturday, February 14, 2009 11:54 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Problem with Schiff's reagent Good Morning! I'm a newcomer to the list, and I'll start by pointing out that I'm not a histologist by any means. I am a freshwater ecologist, and we're trying to study mucus secretion behavior in freshwater snails. Last Spring and last summer, we developed a process whereby we could visualize snail mucus trails on glass slides using a periodic acid-Schiff's reagent staining technique. But now, in following the same protocol as used last year for making our own Schiff's reagent, I cannot get the final solution to filter out clear. Recipe I'm using: 900 ml boiling water 10 grams basic fuchsin 25 ml concentrated HCl acid (12 M) 40 grams sodium metabisulfite (this is essentially Sigma Aldrich's ratios, I think) Let this sit for 24 to 72 hours, take 100 ml aliquot, add 0.75 - 1.0 gram ground activated charcoal, stir for 10 minutes, filter through filter paper then through GF/C glass fiber filter. Last summer I got nice, clear (slightly yellow) and very active Schiff's reagent. But now I cannot seem to get the filtrate to be clear. Even after 10 minutes exposure to ground activated charcoal and filtering, the filtrate remains bright orange to dark red and it does not seem to stain mucus trails very well. All the reagents are the same as those used last summer (i.e., less than 7 months old; although the HCl is a bottle several years old from a different lab). Anyone have any troubleshooting suggestions? I don't know the chemistry very well, but the sodium metabisulfite is used for "decoloring" the initial solution, right? So is the metabisulfite not working for some reason now?? Any help/suggestions would be greatly appreciated. Cheers - Paul Paul E. Brunkow, PhD Department of Biological Sciences Southern Illinois University Edwardsville Edwardsville, IL USA ------------------------------------------------- SIUE Web Mail _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet Confidentiality Notice: This e-mail message, including any attachments, is for the sole use of the intended recipient(s) and may contain confidential and privileged information. Any unauthorized review, use, disclosure or distribution is prohibited. If you are not the intended recipient, please contact the sender by reply e-mail and destroy all copies of the original message. _______________________________________________ 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