Re: [Histonet] Thioflavine S for Amyloid
Congo red is the gold standard for amyloid staining. It is the most sensitive of the amyloid stain, at about 97%. However, sometimes the Congo red will not stain the amyloid protein, such as when the amyloid is a large, very old deposit. In that case, more and more amyloid is being crammed into the same space, and the beta pleats become warped. For Congo red staining to work, the beta pleats must be consistently at a certain distance apart (7 um apart, if I remember correctly). Congo red is a linear dye, with 2 SO3- groups, one at each end. So it binds to the amyloid protein pleats, one dye right after another | | | | |, and that's why it will polarize. But if the beta pleats are not lining up | \ \ | \ / / | | \ /, then the Congo red, may not be able to bind, as the binding sites on the amyloid are not the correct distance apart. And it the Congo red can bind, it is not lining up | | | | |, but is binding in all directions, similar to the warped beta pleats. Therefore it will not birefringe/polarize. Overfixation in formalin will do the same thing, as there will be too many formalin cross-links, warping the beta pleats. Therefore, when it is suspected that the person really has amyloid, but the Congo red isn't working (remember, it's 97% sensitive, which means it's not demonstrating amyloid 3% of the time), it's good to have back ups, which we have used, and have been able to demonstrate amyloid when the Congo red doesn't work. Since the alternatives aren't as specific or sensitive for amyloid as Congo red, when we have to go to our backups, we tend to do all three, on the theory that even though they aren't as sensitive or specific, since they are all staining a different aspect of amyloid, and if all three are showing positivity, then it most be amyloid. Congo red, viewed with fluoresence microscope. Use the auramine-rhodamine AFB filters (hit it with 540 green light), and the Congo red-amyloid will fluoresce orange. Crystal violet or Methyl violet - polychromatic dyes that bind to carboxyl ions on amyloid. So one dye component stains the amyloid a violet color, while the other dye components stain the background a blue-purple. Need to use aqueous mounting media, so not a permanent stain. Not as sensitive (about 70%) or specific as Congo red. Will stain mucin. AL Amyloid also tends to have a lower concentration of surface carboxyl ions, so tends to have negative staining with the CV or MV stains. Thioflavin T (TFT) and Thioflaving S (TFS) are fluorochromes, so need a fluorescence miscroscope, blue light excitation at 490, similar to FITC, and these dyes will fluoresce yellow. These dyes appear to stain the P component on amyloid, which is a pentagonal shaped polysaccharide protein, which is a normal protein in our blood (alpha-globin), but for some reason attaches itself to amyloid. It's a fast stain, easy to do, very sensitive for amyloid. However, you do need a FITC fluoresence microscope, and since it's an aqueous mount, it's not permanent. It's not specific for amyloid, as other components will be stained and fluoresce yellow, or will autofluoresce yellow (such as elastin fibers, dense connective tissue, lipofuchsin, a lot of other granules). I've only used TFT, so I can't say if TFS is any better. So TFS is good for a back up, but I would continue with Congo red, or the Amyloid red from Anatech, to be the primary amyloid stain. Peggy A. Wenk, HTL(ASCP)SLS Beaumont Health Systems Royal Oak, MI 48073 Opinions expressed are mine, and do not reflect on my place of employment. -Original Message- From: Mitchell Jean A Sent: Friday, April 12, 2013 2:20 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Thioflavine S for Amyloid Would appreciated some feedback/input from labs using Thioflavine S staining protocol for amyloid screening. Any advantages/disadvantages to this procedure vs Congo Red? Thanks much!! Jean Mitchell, BS HT (ASCP) University of Wisconsin Hospital Clinics Neuromuscular Laboratory 600 Highland Avenue Madison, WI 53792-5132 ___ 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
Re: [Histonet] Re: Formalin Neutralizer
Benjamin: You are wrong again because formalin is the commercial brand name used to define the about 47% aq. solution of methanal in dist. water and, as a matter of fact, it does contain an additive (methanol) to prevent its polymerization into para-formaldehyde. René J. From: Benjamin benja...@histologistics.com To: Bob Richmond rsrichm...@gmail.com Cc: Histonet@lists.utsouthwestern.edu histonet@lists.utsouthwestern.edu Sent: Saturday, April 13, 2013 6:04 PM Subject: Re: [Histonet] Re: Formalin Neutralizer I am posting that my last comment about formalin was wrong about the odor, it does not have any additives. And as far as dumping it down the drain follow your local regs!! Sent from my iPhone On Apr 13, 2013, at 2:09 PM, Bob Richmond rsrichm...@gmail.com wrote: I'm surprised that the dismal topic of formaldehyde neutralization never seems to get settled. I have a litre of 10% neutral buffered formalin. How many grams of sodium sulfite (or bisulfite, or metabisulfite) do I have to weigh out and pour in to neutralize it? What is the chemical reaction? Is ammonia also required? and if so, I've got the same questions about ammonia. Will the Herrn Inschpektors allow me to use generic chemicals, or do I have to buy some expensive proprietary product in order to satisfy them? Bob Richmond Samurai Pathologist Maryville TN ___ 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 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Amyloid
I think this is a great article to read about amyloid. This is the link to the paper http://propath.com/newsletters-immunohistochemistry-173/181-september-2004-immunohistochemistry-in-amyloidosis It belongs to a great Pathologist Dr. Rodney T. Miller Dr. César RomeroBuenos AiresArgentina ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] RE: California Society for Histotechnology Meeting May 3-5
Can the room block rate be extended? From: James Watson jwat...@gnf.org To: James Watson jwat...@gnf.org, 'Histonet@lists.utsouthwestern.edu' Histonet@lists.utsouthwestern.edu Date: 04/10/2013 07:35 AM Subject:[Histonet] RE: California Society for Histotechnology Meeting May 3-5 Sent by:histonet-boun...@lists.utsouthwestern.edu I was just informed that On Line registration is now working through our website. http://www.californiahistology.org/ I have also been told this can be accessed by smartphone, I-pad. also payment can be made through PayPal or Credit Card. James Watson HT ASCP GNF Genomics Institute of the Novartis Research Foundation Tel858-332-4647 Fax 858-812-1915 jwat...@gnf.org -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [ mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of James Watson Sent: Tuesday, April 09, 2013 7:25 AM To: 'Histonet@lists.utsouthwestern.edu' Subject: [Histonet] California Society for Histotechnology Meeting May 3-5 Joining the other Societies advertising their meeting on the Histonet. Our meeting this year is in Burlingame near the San Francisco international airport. Our annual symposium is about a month away. Hope that you are planning on joining us in Burlingame. Our new website is still having problems with the online registration, our webmaster is working hard to get it going. But we are not sure when it will be available, so you should consider filling out the attached registration from and mailing it to us. We already have 30 Vendors signed up to attend and have outstanding speakers coming from all over the country with a wide variety of backgrounds and subjects. We are hoping to have another great meeting. See you there. Visit our website at to see the program and mail in registration form. http://www.californiahistology.org/ James Watson HT ASCP GNF Genomics Institute of the Novartis Research Foundation Tel858-332-4647 Fax 858-812-1915mailto:858-812-1915jwat...@gnf.org jwat...@gnf.org ___ 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 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] special stain reagents change
Coming from a large reference laboratory, it was crucial to change out reagents for special stains every week since our workload was so heavy. Now at a smaller GI lab, I am wondering if it is absolutely necessary to change out the reagents so frequently? We have changed about 2-3 weeks depending on the amount of slide volume (the kits say up to 300-500 slides- AMT Kits). We were changing every week in the beginning, but one week our reagents were delayed so we changed it out 2 weeks later, and have noticed that the reagents still are fresh and the staining quality is not compromised. The alcohols, blueing, and xylene is changed every day. Any thoughts? Thanks in advance! Sophia ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
