[Histonet] Re: Formalin Fixation issues (Bob Richmond)
If you have large specimens, it is best to open them as soon as possible. If it is a breast, at least do your prelim measurements and slice it open, leaving it still attached, so that the formain can penetrate the tissue. If you go ahead and slice it up, at least wrap it, in order with paper towels, to keep orientation while it fixes. You can always block it and then let it fix too. You just need to be sure to slice it thin so that it fixes; and make sure you have adequate spacing between cassettes. You just need to figure out what way is best for your facility. Amanda Amador, AAS, ASCP(CM) Sr. Histotechnician Pathology Consultants of New Mexico 600 N. Richardson Roswell, NM 88202 575-622-5600, ext 218 From: histonet-boun...@lists.utsouthwestern.edu [histonet-boun...@lists.utsouthwestern.edu] on behalf of histonet-requ...@lists.utsouthwestern.edu [histonet-requ...@lists.utsouthwestern.edu] Sent: Tuesday, July 10, 2012 11:02 AM To: histonet@lists.utsouthwestern.edu Subject: Histonet Digest, Vol 104, Issue 11 Send Histonet mailing list submissions to histonet@lists.utsouthwestern.edu To subscribe or unsubscribe via the World Wide Web, visit http://lists.utsouthwestern.edu/mailman/listinfo/histonet or, via email, send a message with subject or body 'help' to histonet-requ...@lists.utsouthwestern.edu You can reach the person managing the list at histonet-ow...@lists.utsouthwestern.edu When replying, please edit your Subject line so it is more specific than Re: Contents of Histonet digest... Today's Topics: 1. Re: Formalin Fixation issues (Bob Richmond) 2. RE: Need help with CD40 on frozen sections (Teri Johnson) 3. Re: Formalin Fixation issues (Bob Richmond) 4. Day Shift Histotech Job Near Ithaca, NY (Melissa Phelan) 5. Re: Re: Formalin Fixation issues (Jill Cox) 6. RE: Re: Formalin Fixation issues (Weems, Joyce K.) 7. RE: Re: Formalin Fixation issues (Elizabeth Chlipala) 8. GBS IHC (Richard Cartun) 9. AUTO: Ramona Nelson is out of the office. (returning 07/16/2012) (ramona_nel...@bd.com) 10. Subject: Re: [PATHO-L] refreshing perspective on formaldehyde (White, Lisa M.) 11. Beecher tissue array 1 mm. punch set. (Mohammad Sayeeduddin) 12. CD40 (Reynolds,Donna M) 13. IVD P63 (Cathy Crumpton) 14. human vimentin in mouse tissue (Kim Merriam) 15. Re: IVD P63 (Kim Donadio) 16. RE: IVD P63 (Clare Thornton) -- Message: 1 Date: Mon, 9 Jul 2012 13:25:05 -0400 From: Bob Richmond rsrichm...@gmail.com Subject: [Histonet] Re: Formalin Fixation issues To: histonet@lists.utsouthwestern.edu Message-ID: CAOKsRH7dwef0cYYNKuvFjQgxiia_76vHasD-oVwAuk6k=4e...@mail.gmail.com Content-Type: text/plain; charset=ISO-8859-1 Jill Cox asks: Is anyone having problems with breast fixation prior to processing in formalin? For a couple of months now our breast specimens aren't fixing very well before gross. Our pathologist thinks they have changed something in the formalin itself. We utilize 15/1 ratio and in some cases let fix over the weekend. This has only been happening over the last couple of months and can't seem to figure this out. Any advice or similar problems, would love to hear from you. Breast specimens shouldn't be expected to fix before they're grossed, or at least before they've been cut into thin slices. Delaying fixation compromises immunostaining, to say nothing of H E. They should be grossed as promptly as possible after they're received, and should never sit over the weekend without dissection. I seriously doubt that anything in the formalin has changed. It's your technique that needs to change. Unfortunately, this is a difficult task to delegate. Bob Richmond Samurai Pathologist Knoxville TN -- Message: 2 Date: Mon, 9 Jul 2012 17:46:25 + From: Teri Johnson tjohn...@gnf.org Subject: [Histonet] RE: Need help with CD40 on frozen sections To: histonet@lists.utsouthwestern.edu histonet@lists.utsouthwestern.edu Cc: Bea DeBrosse-Serra \(bdebrosse-se...@isisph.com\) bdebrosse-se...@isisph.com Message-ID: 9f3cfee76e51b64991c7485270890b400cdc4...@ex4.lj.gnf.org Content-Type: text/plain; charset=us-ascii Hi Bea! I just left you a voicemail message. I looked up this antibody data sheet and to say the least the staining on the image is underwhelming! I would try fixing the cryosections with Beckstead's zinc prior to staining, and you can even consider fixing the spleens with Zinc and then cryoprotecting with sucrose prior to freezing. It appears they recommend using a biotinylated secondary and then streptavidin-HRP for staining. You might see if that helps as well. Teri Teri Johnson, HT(ASCP)QIHC GNF Histology Lab Manager Genomics Institute of the Novartis Research Foundation 858-332-4752 -- Message: 3
[Histonet] Re: Formalin Fixation issues
Jill Cox asks: Is anyone having problems with breast fixation prior to processing in formalin? For a couple of months now our breast specimens aren't fixing very well before gross. Our pathologist thinks they have changed something in the formalin itself. We utilize 15/1 ratio and in some cases let fix over the weekend. This has only been happening over the last couple of months and can't seem to figure this out. Any advice or similar problems, would love to hear from you. Breast specimens shouldn't be expected to fix before they're grossed, or at least before they've been cut into thin slices. Delaying fixation compromises immunostaining, to say nothing of H E. They should be grossed as promptly as possible after they're received, and should never sit over the weekend without dissection. I seriously doubt that anything in the formalin has changed. It's your technique that needs to change. Unfortunately, this is a difficult task to delegate. Bob Richmond Samurai Pathologist Knoxville TN ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re: Formalin Fixation issues
At this point I'm not sure I understand what Jill Cox's problem is. Forgive me if I keep blaming the pathologist - since I am one! - but in my experience most of these breast problems originate at the gross desk. Is your pathologist cutting the fatty breast tissue thin enough? This is always a challenge, even after you've done it as long as I have. If your pathologist is cramming the cassettes full of fat, then that's where the problem lies. Do you prepare your own neutral buffered formalin, or buy it? I prepared a lot of it years ago, and supervised others who did, and I learned the hard way that mistakes are easy to make when you brew your own. If you buy your NBF ready made, as most people do nowadays, then read the label carefully. Have you changed brands recently? Bob Richmond Samurai Pathologist Knoxville TN *** On Mon, Jul 9, 2012 at 1:38 PM, Jill Cox jco...@yahoo.com wrote: I think you misunderstood what I meant. I know it's not supposed to be fixed before grossing, it's also not getting fixed after processing. But thanks for your two cents.. From: Bob Richmond rsrichm...@gmail.com To: histonet@lists.utsouthwestern.edu Sent: Monday, July 9, 2012 10:25 AM Subject: [Histonet] Re: Formalin Fixation issues Jill Cox asks: Is anyone having problems with breast fixation prior to processing in formalin? For a couple of months now our breast specimens aren't fixing very well before gross. Our pathologist thinks they have changed something in the formalin itself. We utilize 15/1 ratio and in some cases let fix over the weekend. This has only been happening over the last couple of months and can't seem to figure this out. Any advice or similar problems, would love to hear from you. Breast specimens shouldn't be expected to fix before they're grossed, or at least before they've been cut into thin slices. Delaying fixation compromises immunostaining, to say nothing of H E. They should be grossed as promptly as possible after they're received, and should never sit over the weekend without dissection. I seriously doubt that anything in the formalin has changed. It's your technique that needs to change. Unfortunately, this is a difficult task to delegate. Bob Richmond Samurai Pathologist Knoxville TN ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Re: Formalin Fixation issues
Yes Bob, the sections are good and thin. We use Scientific Products, 10% NBF. Just seems like the Formalin is watered down. Going to try a different brand. I have worked for Pathologist's that cram cassettes but this is not the case here. From: Bob Richmond rsrichm...@gmail.com To: histonet@lists.utsouthwestern.edu Sent: Monday, July 9, 2012 10:51 AM Subject: [Histonet] Re: Formalin Fixation issues At this point I'm not sure I understand what Jill Cox's problem is. Forgive me if I keep blaming the pathologist - since I am one! - but in my experience most of these breast problems originate at the gross desk. Is your pathologist cutting the fatty breast tissue thin enough? This is always a challenge, even after you've done it as long as I have. If your pathologist is cramming the cassettes full of fat, then that's where the problem lies. Do you prepare your own neutral buffered formalin, or buy it? I prepared a lot of it years ago, and supervised others who did, and I learned the hard way that mistakes are easy to make when you brew your own. If you buy your NBF ready made, as most people do nowadays, then read the label carefully. Have you changed brands recently? Bob Richmond Samurai Pathologist Knoxville TN *** On Mon, Jul 9, 2012 at 1:38 PM, Jill Cox jco...@yahoo.com wrote: I think you misunderstood what I meant. I know it's not supposed to be fixed before grossing, it's also not getting fixed after processing. But thanks for your two cents.. From: Bob Richmond rsrichm...@gmail.com To: histonet@lists.utsouthwestern.edu Sent: Monday, July 9, 2012 10:25 AM Subject: [Histonet] Re: Formalin Fixation issues Jill Cox asks: Is anyone having problems with breast fixation prior to processing in formalin? For a couple of months now our breast specimens aren't fixing very well before gross. Our pathologist thinks they have changed something in the formalin itself. We utilize 15/1 ratio and in some cases let fix over the weekend. This has only been happening over the last couple of months and can't seem to figure this out. Any advice or similar problems, would love to hear from you. Breast specimens shouldn't be expected to fix before they're grossed, or at least before they've been cut into thin slices. Delaying fixation compromises immunostaining, to say nothing of H E. They should be grossed as promptly as possible after they're received, and should never sit over the weekend without dissection. I seriously doubt that anything in the formalin has changed. It's your technique that needs to change. Unfortunately, this is a difficult task to delegate. Bob Richmond Samurai Pathologist Knoxville 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
RE: [Histonet] Re: Formalin Fixation issues
I would also call the company to see if they have had others with problems. Has it all been the same lot number? Joyce Weems Pathology Manager 678-843-7376 Phone 678-843-7831 Fax joyce.we...@emoryhealthcare.org www.saintjosephsatlanta.org 5665 Peachtree Dunwoody Road Atlanta, GA 30342 This e-mail, including any attachments is the property of Saint Joseph's Hospital and is intended for the sole use of the intended recipient(s). It may contain information that is privileged and confidential. Any unauthorized review, use, disclosure, or distribution is prohibited. If you are not the intended recipient, please delete this message, and reply to the sender regarding the error in a separate email. -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Jill Cox Sent: Monday, July 09, 2012 2:01 PM To: Bob Richmond; Histonet@Lists. Edu Subject: Re: [Histonet] Re: Formalin Fixation issues Yes Bob, the sections are good and thin. We use Scientific Products, 10% NBF. Just seems like the Formalin is watered down. Going to try a different brand. I have worked for Pathologist's that cram cassettes but this is not the case here. From: Bob Richmond rsrichm...@gmail.com To: histonet@lists.utsouthwestern.edu Sent: Monday, July 9, 2012 10:51 AM Subject: [Histonet] Re: Formalin Fixation issues At this point I'm not sure I understand what Jill Cox's problem is. Forgive me if I keep blaming the pathologist - since I am one! - but in my experience most of these breast problems originate at the gross desk. Is your pathologist cutting the fatty breast tissue thin enough? This is always a challenge, even after you've done it as long as I have. If your pathologist is cramming the cassettes full of fat, then that's where the problem lies. Do you prepare your own neutral buffered formalin, or buy it? I prepared a lot of it years ago, and supervised others who did, and I learned the hard way that mistakes are easy to make when you brew your own. If you buy your NBF ready made, as most people do nowadays, then read the label carefully. Have you changed brands recently? Bob Richmond Samurai Pathologist Knoxville TN *** On Mon, Jul 9, 2012 at 1:38 PM, Jill Cox jco...@yahoo.com wrote: I think you misunderstood what I meant. I know it's not supposed to be fixed before grossing, it's also not getting fixed after processing. But thanks for your two cents.. From: Bob Richmond rsrichm...@gmail.com To: histonet@lists.utsouthwestern.edu Sent: Monday, July 9, 2012 10:25 AM Subject: [Histonet] Re: Formalin Fixation issues Jill Cox asks: Is anyone having problems with breast fixation prior to processing in formalin? For a couple of months now our breast specimens aren't fixing very well before gross. Our pathologist thinks they have changed something in the formalin itself. We utilize 15/1 ratio and in some cases let fix over the weekend. This has only been happening over the last couple of months and can't seem to figure this out. Any advice or similar problems, would love to hear from you. Breast specimens shouldn't be expected to fix before they're grossed, or at least before they've been cut into thin slices. Delaying fixation compromises immunostaining, to say nothing of H E. They should be grossed as promptly as possible after they're received, and should never sit over the weekend without dissection. I seriously doubt that anything in the formalin has changed. It's your technique that needs to change. Unfortunately, this is a difficult task to delegate. Bob Richmond Samurai Pathologist Knoxville 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 This e-mail message (including any attachments) is for the sole use of the intended recipient(s) and may contain confidential and privileged information. If the reader of this message is not the intended recipient, you are hereby notified that any dissemination, distribution or copying of this message (including any attachments) is strictly prohibited. If you have received this message in error, please contact the sender by reply e-mail message and destroy all copies of the original message (including attachments). ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
RE: [Histonet] Re: Formalin Fixation issues
There is a real nice article on fixation and handling of breast lumpectomy samples in the Journal of Histotechnology by Dr. Stephen Ruby. It goes over a nice technique we used to use on those samples, worked great. It did use alcoholic formalin which I suspect can't be used currently if you have not validated for that fixative for ER/PR etc. But the overall technique can be used with 10% NBF. The thickness of samples is key as Bob pointed out. We had a new pathologist on gross and the next day we had about 20 or 30 recuts on their breast samples. That number of reacts was uncommon. I needed to figure out what was the issue and we found out it was the size of tissue that was placed into the cassettes. I explained that they needed to place smaller and thinner pieces of tissue in the cassette. Once I did that we did not have anymore problems with their samples. The article information is below. Paper Towel Sandwich - Alcoholic Formalin Fixation for Breast Biopsies Author: Ruby, Stephen G. Source: Journal of Histotechnology, Number 1, March 1999 , pp. 49-51(3) Publisher: Maney Publishing Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC PO Box 18592 Boulder, CO 80308-1592 (303) 682-3949 office (303) 682-9060 fax (303) 881-0763 cell www.premierlab.com Ship to address: 1567 Skyway Drive, Unit E Longmont, CO 80504 -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Bob Richmond Sent: Monday, July 09, 2012 11:51 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Re: Formalin Fixation issues At this point I'm not sure I understand what Jill Cox's problem is. Forgive me if I keep blaming the pathologist - since I am one! - but in my experience most of these breast problems originate at the gross desk. Is your pathologist cutting the fatty breast tissue thin enough? This is always a challenge, even after you've done it as long as I have. If your pathologist is cramming the cassettes full of fat, then that's where the problem lies. Do you prepare your own neutral buffered formalin, or buy it? I prepared a lot of it years ago, and supervised others who did, and I learned the hard way that mistakes are easy to make when you brew your own. If you buy your NBF ready made, as most people do nowadays, then read the label carefully. Have you changed brands recently? Bob Richmond Samurai Pathologist Knoxville TN *** On Mon, Jul 9, 2012 at 1:38 PM, Jill Cox jco...@yahoo.com wrote: I think you misunderstood what I meant. I know it's not supposed to be fixed before grossing, it's also not getting fixed after processing. But thanks for your two cents.. From: Bob Richmond rsrichm...@gmail.com To: histonet@lists.utsouthwestern.edu Sent: Monday, July 9, 2012 10:25 AM Subject: [Histonet] Re: Formalin Fixation issues Jill Cox asks: Is anyone having problems with breast fixation prior to processing in formalin? For a couple of months now our breast specimens aren't fixing very well before gross. Our pathologist thinks they have changed something in the formalin itself. We utilize 15/1 ratio and in some cases let fix over the weekend. This has only been happening over the last couple of months and can't seem to figure this out. Any advice or similar problems, would love to hear from you. Breast specimens shouldn't be expected to fix before they're grossed, or at least before they've been cut into thin slices. Delaying fixation compromises immunostaining, to say nothing of H E. They should be grossed as promptly as possible after they're received, and should never sit over the weekend without dissection. I seriously doubt that anything in the formalin has changed. It's your technique that needs to change. Unfortunately, this is a difficult task to delegate. Bob Richmond Samurai Pathologist Knoxville 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
