RE: [Histonet] Re: Staining on Alcohol Fixed Smears
Thanks for the clarification. I appreciate the info! Tom McNemar, HT(ASCP) Histology Co-ordinator Licking Memorial Health Systems (740) 348-4163 (740) 348-4166 tmcne...@lmhealth.org www.LMHealth.org -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Beth Cox Sent: Monday, December 16, 2013 2:25 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Re: Staining on Alcohol Fixed Smears I agree that with no formalin exposure, there is no crosslinking that needs to be broken, but a then, do nothing approach to antigen retrieval oversimplifies the issue. Alcohol fixation makes changes in the protiens/antibodies also, although different changes than formalin does, and these changes need to be accounted for. In my research on alcohol fixed smears, I found that most antibodies perform best with some antigen retrieval, often gentler than formalin tissues require. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech -- Message: 4 Date: Mon, 16 Dec 2013 11:13:59 -0500 From: Tom McNemartmcne...@lmhealth.org Subject: RE: [Histonet] Re: Staining on Alcohol Fixed Smears To: 'Beth Cox'bethc...@gmail.com, histonet@lists.utsouthwestern.edu histonet@lists.utsouthwestern.edu,levi.fr...@gmail.com levi.fr...@gmail.com Message-ID: E9A90E28259D2F4E84308C5E8EA8F7B4016951E94252@lmhs-exchange Content-Type: text/plain; charset=us-ascii Since there is no exposure to formalin, we do not do antigen retrieval on alcohol fixed specimens. We use the Ventana Benchmark XT and copy the protocol to a new number and use the Wet option rather than paraffin. No retrieval needed. Tom McNemar, HT(ASCP) Histology Co-ordinator Licking Memorial Health Systems (740) 348-4163 (740) 348-4166 tmcne...@lmhealth.org www.LMHealth.org -Original Message- From:histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Beth Cox Sent: Saturday, December 14, 2013 6:30 PM To:histonet@lists.utsouthwestern.edu;levi.fr...@gmail.com Subject: [Histonet] Re: Staining on Alcohol Fixed Smears Levi, I've worked up most of the more common antibodies on alcohol fixed smears. There are two things to remember here: 1) your specimens are alcohol fixed, meaning antigen retrieval needs are different, and 2) they are smears, meaning the cells weren't 'cut open' during microtomy, which impeeds some nuclear staining. Those are two different issues to consider. Some rules of thumb: 1) most cell membrane antigens will stain nicely with minimal changes to the protocols you use for FFPE specimens 2) most cytokeratin antigens stain nicely with little change to your FFPE protocols (if you need to change, start by cutting your antigen retrieval time in half) 3) most nuclear antigens require significant changes from FFPE protocols, and some are essentially impossible to stain using standard antibodies. When I return to work on Monday, I will forward you my protocol 'adjustments' for the antibodies you requested. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech On 12/14/2013 12:00 PM,histonet-requ...@lists.utsouthwestern.edu wrote: Message: 4 Date: Sat, 14 Dec 2013 18:25:08 +0200 From: Levi Friedlevi.fr...@gmail.com Subject: [Histonet] Staining on Alcohol Fixed Smears. To:histonet@lists.utsouthwestern.edu Message-ID: CA+puqxTXL85pEwvR=sqPQA36tMJPm=sx2kv3gyebrbu0he3...@mail.gmail.com Content-Type: text/plain; charset=ISO-8859-1 Hi Everyone. I am looking to stain alcohol fixed smears with a group of antibodies. The antibodies of particular interest are p63, p53 and ki67. Along with these antibodies I am looking for positive nuclear and cytoplastic staining antibodies. If anyone has any experience in staining alcohol fixed slides your information is very appreciated. All the best. -- Sincerely, - Levi Fried ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet This e-mail, including attachments, is intended for the sole use of the individual and/or entity to whom it is addressed, and contains information from Licking Memorial Health Systems which is confidential or privileged. If you are not the intended recipient, nor authorized to receive for the intended recipient, be aware that any disclosure, copying, distribution or use of the contents of this e-mail and attachments is prohibited. If you have received this in error, please advise the sender by reply e-mail and delete the message immediately. You may also contact the LMH Process Improvement Center at 740-348-4641. E-mail transmissions cannot be guaranteed to be secure or error-free as information could be intercepted, corrupted, lost, destroyed, arrive late or incomplete, or contain viruses. The sender therefore does not accept liability for any errors
RE: [Histonet] Re: Staining on Alcohol Fixed Smears
What info do you guys have on acetone fixed smears? Specifically, having trouble getting PAX5 (nuclear) to show good signal without chewing up the cells during antigen retrieval. Susan Foreman, HT (ASCP) KDL Pathology 315 Erin Drive Knoxville, TN 37919 (865)584-1933 -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Tom McNemar Sent: Tuesday, December 17, 2013 5:48 AM To: 'Beth Cox'; histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Re: Staining on Alcohol Fixed Smears Thanks for the clarification. I appreciate the info! Tom McNemar, HT(ASCP) Histology Co-ordinator Licking Memorial Health Systems (740) 348-4163 (740) 348-4166 tmcne...@lmhealth.org www.LMHealth.org -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Beth Cox Sent: Monday, December 16, 2013 2:25 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Re: Staining on Alcohol Fixed Smears I agree that with no formalin exposure, there is no crosslinking that needs to be broken, but a then, do nothing approach to antigen retrieval oversimplifies the issue. Alcohol fixation makes changes in the protiens/antibodies also, although different changes than formalin does, and these changes need to be accounted for. In my research on alcohol fixed smears, I found that most antibodies perform best with some antigen retrieval, often gentler than formalin tissues require. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech -- Message: 4 Date: Mon, 16 Dec 2013 11:13:59 -0500 From: Tom McNemartmcne...@lmhealth.org Subject: RE: [Histonet] Re: Staining on Alcohol Fixed Smears To: 'Beth Cox'bethc...@gmail.com, histonet@lists.utsouthwestern.edu histonet@lists.utsouthwestern.edu,levi.fr...@gmail.com levi.fr...@gmail.com Message-ID: E9A90E28259D2F4E84308C5E8EA8F7B4016951E94252@lmhs-exchange Content-Type: text/plain; charset=us-ascii Since there is no exposure to formalin, we do not do antigen retrieval on alcohol fixed specimens. We use the Ventana Benchmark XT and copy the protocol to a new number and use the Wet option rather than paraffin. No retrieval needed. Tom McNemar, HT(ASCP) Histology Co-ordinator Licking Memorial Health Systems (740) 348-4163 (740) 348-4166 tmcne...@lmhealth.org www.LMHealth.org -Original Message- From:histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Beth Cox Sent: Saturday, December 14, 2013 6:30 PM To:histonet@lists.utsouthwestern.edu;levi.fr...@gmail.com Subject: [Histonet] Re: Staining on Alcohol Fixed Smears Levi, I've worked up most of the more common antibodies on alcohol fixed smears. There are two things to remember here: 1) your specimens are alcohol fixed, meaning antigen retrieval needs are different, and 2) they are smears, meaning the cells weren't 'cut open' during microtomy, which impeeds some nuclear staining. Those are two different issues to consider. Some rules of thumb: 1) most cell membrane antigens will stain nicely with minimal changes to the protocols you use for FFPE specimens 2) most cytokeratin antigens stain nicely with little change to your FFPE protocols (if you need to change, start by cutting your antigen retrieval time in half) 3) most nuclear antigens require significant changes from FFPE protocols, and some are essentially impossible to stain using standard antibodies. When I return to work on Monday, I will forward you my protocol 'adjustments' for the antibodies you requested. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech On 12/14/2013 12:00 PM,histonet-requ...@lists.utsouthwestern.edu wrote: Message: 4 Date: Sat, 14 Dec 2013 18:25:08 +0200 From: Levi Friedlevi.fr...@gmail.com Subject: [Histonet] Staining on Alcohol Fixed Smears. To:histonet@lists.utsouthwestern.edu Message-ID: CA+puqxTXL85pEwvR=sqPQA36tMJPm=sx2kv3gyebrbu0he3...@mail.gmail.com Content-Type: text/plain; charset=ISO-8859-1 Hi Everyone. I am looking to stain alcohol fixed smears with a group of antibodies. The antibodies of particular interest are p63, p53 and ki67. Along with these antibodies I am looking for positive nuclear and cytoplastic staining antibodies. If anyone has any experience in staining alcohol fixed slides your information is very appreciated. All the best. -- Sincerely, - Levi Fried ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet This e-mail, including attachments, is intended for the sole use of the individual and/or entity to whom it is addressed, and contains information from Licking Memorial Health Systems which is confidential or privileged. If you are not the intended recipient, nor authorized to receive for the intended
[Histonet] Re: Staining on Alcohol Fixed Smears
I'm sorry, I have no experience with acetone fixed smears. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech On 12/17/2013 8:40 AM, Susan Foreman wrote: What info do you guys have on acetone fixed smears? Specifically, having trouble getting PAX5 (nuclear) to show good signal without chewing up the cells during antigen retrieval. Susan Foreman, HT (ASCP) KDL Pathology 315 Erin Drive Knoxville, TN 37919 (865)584-1933 -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Tom McNemar Sent: Tuesday, December 17, 2013 5:48 AM To: 'Beth Cox'; histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Re: Staining on Alcohol Fixed Smears Thanks for the clarification. I appreciate the info! Tom McNemar, HT(ASCP) Histology Co-ordinator Licking Memorial Health Systems (740) 348-4163 (740) 348-4166 tmcne...@lmhealth.org www.LMHealth.org -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Beth Cox Sent: Monday, December 16, 2013 2:25 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Re: Staining on Alcohol Fixed Smears I agree that with no formalin exposure, there is no crosslinking that needs to be broken, but a then, do nothing approach to antigen retrieval oversimplifies the issue. Alcohol fixation makes changes in the protiens/antibodies also, although different changes than formalin does, and these changes need to be accounted for. In my research on alcohol fixed smears, I found that most antibodies perform best with some antigen retrieval, often gentler than formalin tissues require. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech -- Message: 4 Date: Mon, 16 Dec 2013 11:13:59 -0500 From: Tom McNemartmcne...@lmhealth.org Subject: RE: [Histonet] Re: Staining on Alcohol Fixed Smears To: 'Beth Cox'bethc...@gmail.com, histonet@lists.utsouthwestern.edu histonet@lists.utsouthwestern.edu,levi.fr...@gmail.com levi.fr...@gmail.com Message-ID: E9A90E28259D2F4E84308C5E8EA8F7B4016951E94252@lmhs-exchange Content-Type: text/plain; charset=us-ascii Since there is no exposure to formalin, we do not do antigen retrieval on alcohol fixed specimens. We use the Ventana Benchmark XT and copy the protocol to a new number and use the Wet option rather than paraffin. No retrieval needed. Tom McNemar, HT(ASCP) Histology Co-ordinator Licking Memorial Health Systems (740) 348-4163 (740) 348-4166 tmcne...@lmhealth.org www.LMHealth.org -Original Message- From:histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Beth Cox Sent: Saturday, December 14, 2013 6:30 PM To:histonet@lists.utsouthwestern.edu;levi.fr...@gmail.com Subject: [Histonet] Re: Staining on Alcohol Fixed Smears Levi, I've worked up most of the more common antibodies on alcohol fixed smears. There are two things to remember here: 1) your specimens are alcohol fixed, meaning antigen retrieval needs are different, and 2) they are smears, meaning the cells weren't 'cut open' during microtomy, which impeeds some nuclear staining. Those are two different issues to consider. Some rules of thumb: 1) most cell membrane antigens will stain nicely with minimal changes to the protocols you use for FFPE specimens 2) most cytokeratin antigens stain nicely with little change to your FFPE protocols (if you need to change, start by cutting your antigen retrieval time in half) 3) most nuclear antigens require significant changes from FFPE protocols, and some are essentially impossible to stain using standard antibodies. When I return to work on Monday, I will forward you my protocol 'adjustments' for the antibodies you requested. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech On 12/14/2013 12:00 PM,histonet-requ...@lists.utsouthwestern.edu wrote: Message: 4 Date: Sat, 14 Dec 2013 18:25:08 +0200 From: Levi Friedlevi.fr...@gmail.com Subject: [Histonet] Staining on Alcohol Fixed Smears. To:histonet@lists.utsouthwestern.edu Message-ID: CA+puqxTXL85pEwvR=sqPQA36tMJPm=sx2kv3gyebrbu0he3...@mail.gmail.com Content-Type: text/plain; charset=ISO-8859-1 Hi Everyone. I am looking to stain alcohol fixed smears with a group of antibodies. The antibodies of particular interest are p63, p53 and ki67. Along with these antibodies I am looking for positive nuclear and cytoplastic staining antibodies. If anyone has any experience in staining alcohol fixed slides your information is very appreciated. All the best. -- Sincerely, - Levi Fried ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet This e-mail, including attachments, is intended for the sole use of the individual and/or entity to whom it is addressed, and contains information from Licking
RE: [Histonet] Re: Staining on Alcohol Fixed Smears
Since there is no exposure to formalin, we do not do antigen retrieval on alcohol fixed specimens. We use the Ventana Benchmark XT and copy the protocol to a new number and use the Wet option rather than paraffin. No retrieval needed. Tom McNemar, HT(ASCP) Histology Co-ordinator Licking Memorial Health Systems (740) 348-4163 (740) 348-4166 tmcne...@lmhealth.org www.LMHealth.org -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Beth Cox Sent: Saturday, December 14, 2013 6:30 PM To: histonet@lists.utsouthwestern.edu; levi.fr...@gmail.com Subject: [Histonet] Re: Staining on Alcohol Fixed Smears Levi, I've worked up most of the more common antibodies on alcohol fixed smears. There are two things to remember here: 1) your specimens are alcohol fixed, meaning antigen retrieval needs are different, and 2) they are smears, meaning the cells weren't 'cut open' during microtomy, which impeeds some nuclear staining. Those are two different issues to consider. Some rules of thumb: 1) most cell membrane antigens will stain nicely with minimal changes to the protocols you use for FFPE specimens 2) most cytokeratin antigens stain nicely with little change to your FFPE protocols (if you need to change, start by cutting your antigen retrieval time in half) 3) most nuclear antigens require significant changes from FFPE protocols, and some are essentially impossible to stain using standard antibodies. When I return to work on Monday, I will forward you my protocol 'adjustments' for the antibodies you requested. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech On 12/14/2013 12:00 PM, histonet-requ...@lists.utsouthwestern.edu wrote: Message: 4 Date: Sat, 14 Dec 2013 18:25:08 +0200 From: Levi Friedlevi.fr...@gmail.com Subject: [Histonet] Staining on Alcohol Fixed Smears. To:histonet@lists.utsouthwestern.edu Message-ID: CA+puqxTXL85pEwvR=sqPQA36tMJPm=sx2kv3gyebrbu0he3...@mail.gmail.com Content-Type: text/plain; charset=ISO-8859-1 Hi Everyone. I am looking to stain alcohol fixed smears with a group of antibodies. The antibodies of particular interest are p63, p53 and ki67. Along with these antibodies I am looking for positive nuclear and cytoplastic staining antibodies. If anyone has any experience in staining alcohol fixed slides your information is very appreciated. All the best. -- Sincerely, - Levi Fried ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet This e-mail, including attachments, is intended for the sole use of the individual and/or entity to whom it is addressed, and contains information from Licking Memorial Health Systems which is confidential or privileged. If you are not the intended recipient, nor authorized to receive for the intended recipient, be aware that any disclosure, copying, distribution or use of the contents of this e-mail and attachments is prohibited. If you have received this in error, please advise the sender by reply e-mail and delete the message immediately. You may also contact the LMH Process Improvement Center at 740-348-4641. E-mail transmissions cannot be guaranteed to be secure or error-free as information could be intercepted, corrupted, lost, destroyed, arrive late or incomplete, or contain viruses. The sender therefore does not accept liability for any errors or omissions in the contents of this message, which arise as a result of e-mail transmission. Thank you. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re: Staining on Alcohol Fixed Smears
I agree that with no formalin exposure, there is no crosslinking that needs to be broken, but a then, do nothing approach to antigen retrieval oversimplifies the issue. Alcohol fixation makes changes in the protiens/antibodies also, although different changes than formalin does, and these changes need to be accounted for. In my research on alcohol fixed smears, I found that most antibodies perform best with some antigen retrieval, often gentler than formalin tissues require. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech -- Message: 4 Date: Mon, 16 Dec 2013 11:13:59 -0500 From: Tom McNemartmcne...@lmhealth.org Subject: RE: [Histonet] Re: Staining on Alcohol Fixed Smears To: 'Beth Cox'bethc...@gmail.com, histonet@lists.utsouthwestern.edu histonet@lists.utsouthwestern.edu,levi.fr...@gmail.com levi.fr...@gmail.com Message-ID: E9A90E28259D2F4E84308C5E8EA8F7B4016951E94252@lmhs-exchange Content-Type: text/plain; charset=us-ascii Since there is no exposure to formalin, we do not do antigen retrieval on alcohol fixed specimens. We use the Ventana Benchmark XT and copy the protocol to a new number and use the Wet option rather than paraffin. No retrieval needed. Tom McNemar, HT(ASCP) Histology Co-ordinator Licking Memorial Health Systems (740) 348-4163 (740) 348-4166 tmcne...@lmhealth.org www.LMHealth.org -Original Message- From:histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Beth Cox Sent: Saturday, December 14, 2013 6:30 PM To:histonet@lists.utsouthwestern.edu;levi.fr...@gmail.com Subject: [Histonet] Re: Staining on Alcohol Fixed Smears Levi, I've worked up most of the more common antibodies on alcohol fixed smears. There are two things to remember here: 1) your specimens are alcohol fixed, meaning antigen retrieval needs are different, and 2) they are smears, meaning the cells weren't 'cut open' during microtomy, which impeeds some nuclear staining. Those are two different issues to consider. Some rules of thumb: 1) most cell membrane antigens will stain nicely with minimal changes to the protocols you use for FFPE specimens 2) most cytokeratin antigens stain nicely with little change to your FFPE protocols (if you need to change, start by cutting your antigen retrieval time in half) 3) most nuclear antigens require significant changes from FFPE protocols, and some are essentially impossible to stain using standard antibodies. When I return to work on Monday, I will forward you my protocol 'adjustments' for the antibodies you requested. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech On 12/14/2013 12:00 PM,histonet-requ...@lists.utsouthwestern.edu wrote: Message: 4 Date: Sat, 14 Dec 2013 18:25:08 +0200 From: Levi Friedlevi.fr...@gmail.com Subject: [Histonet] Staining on Alcohol Fixed Smears. To:histonet@lists.utsouthwestern.edu Message-ID: CA+puqxTXL85pEwvR=sqPQA36tMJPm=sx2kv3gyebrbu0he3...@mail.gmail.com Content-Type: text/plain; charset=ISO-8859-1 Hi Everyone. I am looking to stain alcohol fixed smears with a group of antibodies. The antibodies of particular interest are p63, p53 and ki67. Along with these antibodies I am looking for positive nuclear and cytoplastic staining antibodies. If anyone has any experience in staining alcohol fixed slides your information is very appreciated. All the best. -- Sincerely, - Levi Fried ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re: Staining on Alcohol Fixed Smears
Levi, I've worked up most of the more common antibodies on alcohol fixed smears. There are two things to remember here: 1) your specimens are alcohol fixed, meaning antigen retrieval needs are different, and 2) they are smears, meaning the cells weren't 'cut open' during microtomy, which impeeds some nuclear staining. Those are two different issues to consider. Some rules of thumb: 1) most cell membrane antigens will stain nicely with minimal changes to the protocols you use for FFPE specimens 2) most cytokeratin antigens stain nicely with little change to your FFPE protocols (if you need to change, start by cutting your antigen retrieval time in half) 3) most nuclear antigens require significant changes from FFPE protocols, and some are essentially impossible to stain using standard antibodies. When I return to work on Monday, I will forward you my protocol 'adjustments' for the antibodies you requested. Beth Cox, HTL/SCT(ASCP)QIHC Vagabond Histotech On 12/14/2013 12:00 PM, histonet-requ...@lists.utsouthwestern.edu wrote: Message: 4 Date: Sat, 14 Dec 2013 18:25:08 +0200 From: Levi Friedlevi.fr...@gmail.com Subject: [Histonet] Staining on Alcohol Fixed Smears. To:histonet@lists.utsouthwestern.edu Message-ID: CA+puqxTXL85pEwvR=sqPQA36tMJPm=sx2kv3gyebrbu0he3...@mail.gmail.com Content-Type: text/plain; charset=ISO-8859-1 Hi Everyone. I am looking to stain alcohol fixed smears with a group of antibodies. The antibodies of particular interest are p63, p53 and ki67. Along with these antibodies I am looking for positive nuclear and cytoplastic staining antibodies. If anyone has any experience in staining alcohol fixed slides your information is very appreciated. All the best. -- Sincerely, - Levi Fried ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
