Hi Kim and Patsy, This protocol works well on EDTA decalcified FFPE tissue samples.
Tartrate-Resistant Acid Phosphatase (TRAP) histochemistry For Paraffin: Deparaffinize sections through xylenes and alcohol and bring to running deionized water for 5 minutes. For Plastic: Deplasticize through three 20-minute changes of 2-methoxyethyl acetate, two 5-minute changes of acetone followed by running deionized water for 5 minutes. Make up acetate buffer in the volume needed for the number of slides to be stained. 0.2 M ACETATE BUFFER Distilled water for 50.0 ml for 200.0 ml 0.2 M sodium acetate (FW 82.03) 0.82 gm 3.28 gm 50 mM L(+) tartaric acid (FW 230.1) 0.58 gm 2.32 gm Stir using a magnetic stir bar until dissolved; pH to 5.0 Following the 5-minute wash under running water, incubate the sections in 0.2 M acetate buffer for 20 minutes at room temperature. After this time has elapsed, to this same acetate buffer add: for 50.0 ml for 200.0 ml 0.5 mg/ml naphtol AS-MX phosphate 25.0 mg 100.0 mg 1.1 mg/ml fast red TR salt 55.0 mg 220.0 mg OR Add the naphtol AS-MX phosphate and TR salt to freshly prepared and preheated 0.2 M sodium acetate buffer Incubate the sections from 1 – 4 hours at 37C, monitoring after the first hour, until osteoclasts are bright red. Rinse in distilled water, followed by counterstaining in Mayer’s hematoxylin for 1 – 5 minutes (depending on the age of the hematoxylin), wash in running water for 5 minutes, air-dry and mount with permount. Ref. Erlebacher & Derynck J Cell Biol 132: 195 – 210, 1996 September 30, 2003 (Revised) Hermina Borgerink Wake Forest University Health Sciences -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Patsy Ruegg Sent: Monday, February 21, 2011 12:46 PM To: 'Kim Merriam'; 'Histonet' Subject: RE: SPAM-LOW: [Histonet] TRAP staining kit Kim if you are asking if TRAP enzyme histochemistry works on formalin fixed paraffin embedded decalcified bone in my experience it does not. It works on smears and cytology slides or GMA embedded bone without decal, it may work on ETA decaled bone but I do not think it works on ffpe slides. There is an antibody to TRAP now you could use on ffpe sections by IHC. What are you trying to use it for, osteoclasts??? Regards, Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech 12635 Montview Blvd. Ste.215 Aurora, CO 80045 720-859-4060 fax 720-859-4110 www.ihctech.net www.ihcrg.org -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Kim Merriam Sent: Monday, February 21, 2011 7:39 AM To: Histonet Subject: SPAM-LOW: [Histonet] TRAP staining kit Hi All, Does anyone know if there is a TRAP staining kit available for histology (and who sells it)? I have found a bunch of them on google, but they are all for cell culture. Thanks, Kim Kim Merriam, MA, HT(ASCP)QIHC Cambridge, MA _______________________________________________ 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
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