Mohamed,
The University of Nottingham Histology department has an extensive website of histology protocols: http://www.nottingham.ac.uk/pathology/default.html There is a protocol for PAS/OG on that site. I do not have any experience using it, but as it comes out of John Bancroft's old lab it will undoubtedly be very sound. It does differ somewhat from the method of AGE Pearse that I followed many, many moons ago in Glasgow. We also always performed an OFG (orange-fuchsin-green) method on all pituitary bxs (if you need a copy of that let me know).As I'm sure you are aware, tinctorial stains for pituitary adenomas have been replaced with immunohistochemistry for the hormones ACTH, FSH, GH, LH, prolactin and TSH: PAS/OG 1. Section to 70% alcohol 2. leave in 70% alcohol for 5 minutes 3. transfer to Solution "A" for 15 minutes at room temp 4. Wash in 70% alcohol for 5 minutes 5. transfer to reducing rinse "B" for 5 minutes 6. 70% alcohol for 3 minutes 7. rinse in running tap water 8. Schiff's reagent 20-30 minutes 9. wash in water 3 minutes 10. Celestin Blue solution 10 minutes 11. Rinse well in water 12. Hematoxylin 10 minutes 13. wash in water 14. 2% Orange G in 5% phosphotungstic acid for 3 minutes 15. wash in water 16. dehydrate, clear and mount Result Nuclei - black RBC's - yellow-orange Alpha granules - orange Beta granules - magenta (PAS +ve) Solution "A" Periodic acid - 400mg Distilled water - 15 ml 100% alcohol - 35 ml Reducing Solution "B" Sodium thiosulphate - 1gm Distilled water - 20ml 100% alcohol - 30ml HCl - 0.5ml Rationale PAS converts 1,2 glycols to aldehyde. The reducing step is thought to block the subsequent reaction of collagen and retic fibers with the Schiff's reagent. Beta granules contain muco- or glycoproteins that react with Schiff's to give their magenta staining. Orange G, which is an acidic dye, stains the alpha granules. As RBC's are also acidophilic in nature, they also pick up the orange coloration. Reference Pearse AGE. The cytochemical demonstration of gonadotrophic hormone in the human anterior hypophysis. J Pathol Bacteriol. 1949: 61; 195-202 If you need a recipe for the celestin blue solution, drop me a line and I'll get it to you, but you should find it in most good histotechnology text-books, or on-line Best wishes Ronnie Ronnie Houston, MS HT(ASCP)QIHC Anatomic Pathology Manager ChildLab, a Division of Nationwide Children's Hospital www.childlab.com 700 Children's Drive Columbus, OH 43205 (P) 614-722-5450 (F) 614-722-2899 ronald.hous...@nationwidechildrens.org<mailto:ronald.hous...@nationwidechildrens.org> www.NationwideChildrens.org<http://www.NationwideChildrens.org> -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of mohamed abd el razik Sent: Friday, October 22, 2010 8:36 AM To: Histonet@lists.utsouthwestern.edu Subject: [Histonet] pituitary stain --- dear all can you share me pituitary stain protocol called PAS/ celestine blue / Orange G? thanks in advance mohamed _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ----------------------------------------- Confidentiality Notice: The following mail message, including any attachments, is for the sole use of the intended recipient(s) and may contain confidential and privileged information. The recipient is responsible to maintain the confidentiality of this information and to use the information only for authorized purposes. If you are not the intended recipient (or authorized to receive information for the intended recipient), you are hereby notified that any review, use, disclosure, distribution, copying, printing, or action taken in reliance on the contents of this e-mail is strictly prohibited. If you have received this communication in error, please notify us immediately by reply e-mail and destroy all copies of the original message. Thank you. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet