Re: [Histonet] Nitroblue Tetrazolium Chloride
I used to do a stain on post mortem heart slices to look for recent infarctions. the entire tissue turned a sort of bluey colour which remained through formain fixation and processing. the tissue however was incubated in solution of NBT in a sodium cyanide containing buffer - not sure if that would still be allowed these days - but if you want - i could send you the protocol we used best regards On Thu, Oct 28, 2010 at 8:51 PM, Laurie Colbert laurie.colb...@huntingtonhospital.com wrote: I previously posted a question regarding Nitroblue Tetrazolium Chloride, but I didn't really receive the info that I needed - so I thought I would ask again. I need to purchase this item for a research doc. He wants to immerse tissue in this solution for 24 hours and then process as usual. It is my understanding that this is some kind of dye. I see that I can order it from Sigma in tablet form. I've seen it from other companies in a powder form. Has anyone ever used this reagent in the capacity that I describe? Is it available as a ready-to-use solution/liquid? Is there a certain strength/percentage that I should use? Thanks, Laurie Colbert ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet -- Louise Renton Bone Research Unit University of the Witwatersrand Johannesburg South Africa +27 11 717 2298 (tel fax) 073 5574456 (emergencies only) There are nights when the wolves are silent and only the moon howls. George Carlin No trees were killed in the sending of this message. However, many electrons were terribly inconvenienced. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
RE: [Histonet] RE: breast fixation times
One more question regarding ER/PR/Her2..who am I kidding, we will be talking about them forever!! With regards to the ER/PR publication: The PR clone (Ventana 1E2) that we currently use is not listed as 'acceptable'. Is anyone else in this situation? Are you switching to another clone?? Have you come across anything speaking of this scenario? What to do if you currently run a clone not mentioned? It will be a relatively easy validation for me, but I can't image I am alone here. Thanks, Melissa -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Feher, Stephen Sent: Thursday, October 28, 2010 5:50 PM To: Tench, Bill; Weems, Joyce; histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] RE: breast fixation times Great discussion, comprehensive yet concise. Thanks Bill and Joyce and Melissa. Steve -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Tench, Bill Sent: Wednesday, October 27, 2010 12:26 PM To: Weems, Joyce; histonet@lists.utsouthwestern.edu Subject: [Histonet] RE: breast fixation times My apologies for not including the updates accurate for ER and PR. Bill Tench Associate Dir. Laboratory Services Chief, Cytology Services Palomar Medical Center 555 E. Valley Parkway Escondido, California 92025 bill.te...@pph.org Voice: 760- 739-3037 Fax: 760-739-2604 -Original Message- From: Weems, Joyce [mailto:jwe...@sjha.org] Sent: Wednesday, October 27, 2010 9:22 AM To: Tench, Bill; histonet@lists.utsouthwestern.edu Subject: RE: breast fixation times Thanks for this good explanation, Bill. One can not follow the guidelines and document the variant in the report, but not following them could hurt the patient if there is a clinical trial they might participate in. Clinical trials follow the protocol to the letter and if the FDA requirement is not met, the patient can not participate. The times were extended for ER and PR to 72 hours, but NOT yet for Her2. So...because the tissue is all the same, we must follow the 48 hour limit. We just had a case this weekend. Had the clinical staff remove it from the processor on Sun morning and embedded it Monday. We don't ususally have this problem as we are a 6-day lab, but it was finished too late on Fri. Cheers,j Joyce Weems Pathology Manager Saint Joseph's Hospital 5665 Peachtree Dunwoody Rd NE Atlanta, GA 30342 678-843-7376 - Phone 678-843-7831 - Fax -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Tench, Bill Sent: Wednesday, October 27, 2010 11:50 To: histonet@lists.utsouthwestern.edu Subject: [Histonet] breast fixation times There is no exception for core biopsies, as reasonable as that may seem. I have had that discussion with the purveyors of the guidelines. 6-48 is the current standard. there was a lot of discussion about exceeding 48 and using the FISH option. My colleague responsible for this wrote: It is in the CAP checklist, ANP 22998: If the laboratory assesses Her2 by IHC or Her2 gene amplification by in-situ hybridization (FISH, CISH, SISH), does the lab have a documented procedure for ensuring appropriate length of fixation of specimens tested? Specimens subject to Her2 testing should be fixed in 10% neutral buffered formalin for at least 6 hours and no longer than 48 hours. While fixation outside of these time limits is not an absolute exclusion criterion for Her2 testing, labs should qualify any negative results for specimens fixed less than 6 hours or longer than 48 hours. For cases with negative results by IHC, consideration should be given to performing confirmatory analysis by in-situ hybridization. There is also a table in the original ASCO/CAP Guideline Recommendations for Her2 in Breast Cancer (Arch Pathol Lab Med, Vol 131, Jan 2007) that states that tissue fixed in formalin for greater than 48 hours is not an absolute exclusion criterion, but if known to be fixed longer than 48 hours or unknown, the report should qualify any negative result with this information (table 6). As for upcoming changes, i don't know other than these time limitations are suppose to be more rigorously applied to ER and PR, along with the newly instituted documentation of time between excision and time placed in fixative. Bill Tench Associate Dir. Laboratory Services Chief, Cytology Services Palomar Medical Center 555 E. Valley Parkway Escondido, California 92025 bill.te...@pph.org Voice: 760- 739-3037 Fax: 760-739-2604 [None] made the following annotations - Confidential E-Mail: This e-mail is intended only for the person or entity to which it is addressed, and may contain information that is privileged, confidential, or otherwise protected from disclosure. Dissemination, distribution, or
RE: [Histonet] RE: Saponin Technique
for removing erythrocytes from hemorrhagic cytology samples we use filtration of bloody suspension through membrane filter with pore size 20 mikrons, you can find an article in Cytopathology. 2007 Jun;18(3):175-9. Epub 2007 Mar 27. it is worth trying! regards Irena From: tmcne...@lmhealth.org To: valerie.han...@parrishmed.com; histonet@lists.utsouthwestern.edu Date: Mon, 25 Oct 2010 11:52:31 -0400 CC: Subject: [Histonet] RE: Saponin Technique I don't know about the procedure you have but ours only uses a 1% solution of saponin, a 3% solution of calcium gluconate, and a balanced salt solution. We get the balanced salt solution from our pharmacy. 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 Hannen, Valerie Sent: Monday, October 25, 2010 11:40 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] FW: Saponin Technique From: Hannen, Valerie Sent: Monday, October 25, 2010 11:39 AM To: 'histo...@list.utsouthwestern.edu' Subject: Saponin Technique Hi folks.. I am hoping someone out there in Histo-land can help. One of our Pathologists is asking us to check into doing the Saponin Technique to lyse the red cells in some of our cytology specimens. She gave a copy of one of the pages from one of her manuals, and it shows that in order for us to do this we would need to buy alot of reagents that we currently do not have. Being that we would probably not be doing this technique very often, and not sure how cost effective it will be for us to buy the reagents, we are wondering if anyone knows of a commercial product that we can buy that would be a cheaper way out. Thanks in advance!! Valerie Hannen, Histotechnologist Parrish Medical Center Titusville,Florida ** This email is intended solely for the use of the individual to whom it is addressed and may contain information that is privileged, confidential or otherwise exempt from disclosure under applicable law. If the reader of this email is not the intended recipient or the employee or agent responsible for delivering the message to the intended recipient, you are hereby notified that any dissemination, distribution, or copying of this communication is strictly prohibited. If you have received this communication in error, please immediately delete this message. Thank you ** ___ 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 mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Nashville HT Position
Great opportunity for a Histotechnician in a brand new laboratory! Bellmeade Dermatology in Nashville, TN is looking for a certified HT or HTL to run their newly constructed laboratory. Bellmeade Dermatology has been in the dermatology business for 18 years with 3 physicians and 2 Nurse Practitioners' . Candidate must be ASCP certified and CLIA certified to perform gross dissection, prior supervisory experience preferred. The candidate will be responsible for the following: Creation and maintenance of policies and procedures to CLIA standards, leading lab through CLIA inspection, maintenance and quality control for equipment, and routine histology duties. This is a part time position that offers a competitive salary and the flexible hours allows you to put your own personal stamp on the laboratory . Interested applicants should contact Meredith Hale phone 214-596-2219 or through email mh...@carisls.com *** Please no recruiter's or vendors , HT applicant's only , thank you . Meredith Hale HT (ASCP) CM Operations Liaison Director and Education Coordinator Caris Life Sciences 6655 North MacArthur Blvd, Irving Texas 75039 direct: 214-596-2219 cell: 469-648-8253 fax: 972-929-9966 mh...@carisls.com ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] RE: Nitroblue Tetrazolium Chloride
Nitro BT is a tetrazolium salt used in many dehydrogenase enzyme histochemical techniques. It is used in powder form (I would recommend the Nitrotetrazolium Blue chloride from Sigma, cat N6876). The Nitro BT test is used for the demonstration of early myocardial infarction in gross hearts at necropsy by a dehydrogenase enzyme histochemical reaction on slices on unfixed heart. In brief, a redox indicator is reduced by mitochondrial enzymes in cardiac muscle cells producing a blue staining reaction in all but infarcted areas. The loss of enzyme activity from the tissue defines the damaged heart muscle Incubation Medium: Nitro BT50mg 0.2M Tris-HCl buffer, pH 7.4100ml Sodium cyanide 50mg β-NAD 100mg Adjust pH to 7.1 The stock solution can be kept for several weeks at 4C Protocol: 1.Rinse slices of myocardium in cold running water to remove the blood. 2.Incubate slices at 37C for 30 minutes 3.Wash in running water 4.Store in formol saline Result: Normal myocardium - dark blue Areas of infarcted muscle show diminished staining, the degree of which depends on age of infarct Notes: Loss of staining can be detected as early as 1-5 hr after clinically determined onset of infarct. Material can be tested for 3 days after death and even longer if stored at 4C Cyanide is added to the incubation medium to bind keto acids, produced during the dehydrogenation of some hydroxy acids, which may inhibit the dehydrogenase reaction. NAD is added to the reaction medium as it is lost from the tissue rapidly post-mortem, and greatly improves the sensitivity of the reaction. Some reaction medium call for the addition of the electron transfer mediator phenazine methosulfate (PMS), ostensibly to increase the reaction velocity of the enzyme reaction. This is used to by-pass the reaction of tetrazolium reductases, mediating between the reduced coenzymes and the tetrazolium salt. However, if PMS is used, any active cytochrome oxidase must be inhibited, as it can either oxidize the PMS reducing the amount of formazan deposited at the site of the enzyme reaction or even reverse the pattern producing false non-selective staining and obscuring areas of recent myocardial damage. 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.orgmailto:ronald.hous...@nationwidechildrens.org www.NationwideChildrens.orghttp://www.NationwideChildrens.org One person with passion is better than forty people merely interested. ~ E.M. Forster -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Laurie Colbert Sent: Thursday, October 28, 2010 2:52 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Nitroblue Tetrazolium Chloride I previously posted a question regarding Nitroblue Tetrazolium Chloride, but I didn't really receive the info that I needed - so I thought I would ask again. I need to purchase this item for a research doc. He wants to immerse tissue in this solution for 24 hours and then process as usual. It is my understanding that this is some kind of dye. I see that I can order it from Sigma in tablet form. I've seen it from other companies in a powder form. Has anyone ever used this reagent in the capacity that I describe? Is it available as a ready-to-use solution/liquid? Is there a certain strength/percentage that I should use? Thanks, Laurie Colbert ___ 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
RE: [Histonet] lymph nodes peeling out of OCT
I place all tissues in liquid OCT for at least 15 minutes before freezing. This virtually eliminates the problem of tissue sections separating from the OCT after sectioning. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Precipitating Silver
Has anyone heard of precipitating silver out of a solution by adding table salt? I read this somewhere as a means of disposing of silver easily. Any info would be appreciated. Thanks, Sheila Haas Laboratory Supervisor MicroPath Laboratories, Inc. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] PLP fixative
I am seeking basic information on PLP fixative. What is it? Can I make it or buy it? How long does fixation take for lymph node? Thanks in advance, Tom Truscott ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
RE: [Histonet] Precipitating Silver
Yes, it is easy to do. Since silver chloride is insoluble in water, chloride added to a solution containing ionic silver will cause the white compound to precipitate out immediately. It will settle out onto the bottom of the container overnight, or can be filtered out. -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Sheila Haas Sent: Friday, October 29, 2010 1:56 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Precipitating Silver Has anyone heard of precipitating silver out of a solution by adding table salt? I read this somewhere as a means of disposing of silver easily. Any info would be appreciated. Thanks, Sheila Haas Laboratory Supervisor MicroPath Laboratories, Inc. ___ 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] PLP fixative
Follow the links for a receipe. www.pathology.ufl.edu/~molecular/PLP%20Fixative.doc http://www.niehs.nih.gov/research/atniehs/labs/lep/path-support/immuno/reagents.cfm www.hopkinsmedicine.org/.../multimedia/text_documents/Recipes_For_Making_PLP_Fixative.doc William DeSalvo, B.S., HTL(ASCP) From: ttrus...@vetmed.wsu.edu To: histonet@lists.utsouthwestern.edu Date: Fri, 29 Oct 2010 10:55:45 -0700 Subject: [Histonet] PLP fixative I am seeking basic information on PLP fixative. What is it? Can I make it or buy it? How long does fixation take for lymph node? Thanks in advance, Tom Truscott ___ 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] IHC Staining Problem
I hope someone can help me with a problem I've been having with my CD79a hand stain. This is a stain that was validated and working perfectly, when a few months ago I started having reduced, to no staining on my controls, with the occasional run staining a little stronger. I followed the discussion not too long ago with the person who was having similar problems, but the suggestions didn't seem to apply (I use Tween 20 in my PBS rinses, and we use APEX slides from Surgipath-not Fischer Plus). I've tried everything I can think of and I hope someone may have some other ideas. For reference this is for a veterinary diagnostic lab, not a human lab, and this is the only stain we've been having this issue with. My protocol is as follows: After deparfinization/rehydration, AR for 20 min in the steamer in Biogenex Citra Solution (I've gotten new Citra Solution, tried a pressure cooker) 5 min peroxide quench, followed by a 10 min protein block using Dako's Protein Block-Serum Free 60 min Primary incubation (Dako CD79a) at 37 C (decreased dilution from 1:100 to 1:65, new bottle of antibody) 30 min incubation at room temp of ImmPRESS anti-Mouse Reagent from Vector Develop stain with DAB chromogen from Vector Washes of PBS/Tween 20 follow between steps Thank you, Jennifer Hill Research Assistant Kansas State University Veterinary Diagnostic Lab ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Laura Miller is Out of the Office.
I will be out of the office starting 10/29/2010 and will not return until 11/01/2010. I am out of the office for the rest of the day. I will be back on Monday! __ This email has been scanned by the MessageLabs Email Security System. For more information please visit http://www.messagelabs.com/email __ ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
RE: [Histonet] RE: breast fixation times
Melissa, Just because the clone you are using is not recommended in the guidelines does not mean that you cannot use it. You have to validate it the same as using a clone they do recommend, but the bottom line is you can use what ever you want in what ever way you want as long as you validate the protocol on samples generated in your lab. If you want to use a fixation or processing different from what is recommended you have to compare it to formalin fixation and standard (whatever that is) paraffin processing, you would have to do a side by side comparison on the same tissue. As long as you follow the fixation and processing guidelines you can use the ab u want to use without comparing it to abs they recommend, as far as I can tell. You must validate that ab on at least 25 samples generated in your institution just as you would have to validate an antibody they do recommend. 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 Kuhnla, Melissa Sent: Friday, October 29, 2010 3:51 AM To: Feher, Stephen; Tench, Bill; Weems, Joyce; histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] RE: breast fixation times One more question regarding ER/PR/Her2..who am I kidding, we will be talking about them forever!! With regards to the ER/PR publication: The PR clone (Ventana 1E2) that we currently use is not listed as 'acceptable'. Is anyone else in this situation? Are you switching to another clone?? Have you come across anything speaking of this scenario? What to do if you currently run a clone not mentioned? It will be a relatively easy validation for me, but I can't image I am alone here. Thanks, Melissa -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Feher, Stephen Sent: Thursday, October 28, 2010 5:50 PM To: Tench, Bill; Weems, Joyce; histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] RE: breast fixation times Great discussion, comprehensive yet concise. Thanks Bill and Joyce and Melissa. Steve -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Tench, Bill Sent: Wednesday, October 27, 2010 12:26 PM To: Weems, Joyce; histonet@lists.utsouthwestern.edu Subject: [Histonet] RE: breast fixation times My apologies for not including the updates accurate for ER and PR. Bill Tench Associate Dir. Laboratory Services Chief, Cytology Services Palomar Medical Center 555 E. Valley Parkway Escondido, California 92025 bill.te...@pph.org Voice: 760- 739-3037 Fax: 760-739-2604 -Original Message- From: Weems, Joyce [mailto:jwe...@sjha.org] Sent: Wednesday, October 27, 2010 9:22 AM To: Tench, Bill; histonet@lists.utsouthwestern.edu Subject: RE: breast fixation times Thanks for this good explanation, Bill. One can not follow the guidelines and document the variant in the report, but not following them could hurt the patient if there is a clinical trial they might participate in. Clinical trials follow the protocol to the letter and if the FDA requirement is not met, the patient can not participate. The times were extended for ER and PR to 72 hours, but NOT yet for Her2. So...because the tissue is all the same, we must follow the 48 hour limit. We just had a case this weekend. Had the clinical staff remove it from the processor on Sun morning and embedded it Monday. We don't ususally have this problem as we are a 6-day lab, but it was finished too late on Fri. Cheers,j Joyce Weems Pathology Manager Saint Joseph's Hospital 5665 Peachtree Dunwoody Rd NE Atlanta, GA 30342 678-843-7376 - Phone 678-843-7831 - Fax -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Tench, Bill Sent: Wednesday, October 27, 2010 11:50 To: histonet@lists.utsouthwestern.edu Subject: [Histonet] breast fixation times There is no exception for core biopsies, as reasonable as that may seem. I have had that discussion with the purveyors of the guidelines. 6-48 is the current standard. there was a lot of discussion about exceeding 48 and using the FISH option. My colleague responsible for this wrote: It is in the CAP checklist, ANP 22998: If the laboratory assesses Her2 by IHC or Her2 gene amplification by in-situ hybridization (FISH, CISH, SISH), does the lab have a documented procedure for ensuring appropriate length of fixation of specimens tested? Specimens subject to Her2 testing should be fixed in 10% neutral buffered formalin for at least 6 hours and no longer than 48 hours. While fixation outside of these time limits is not an absolute exclusion criterion for Her2 testing, labs should qualify any
[Histonet] cryostat question
Does anyone have 'hands on' experience performing Mohs using a countertop cryostat? Can the specimen head be manipulated to acccomodate a badly embedded block? Does the machine stay cold enough to perform Mohs? Any advice would be appreciated. My e-mail is histo...@aol.com. Thanks, Susan ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] IHC Staining Problem
How far in advance are your controls being cut? Are your patient tissues staining? Jennifer Hill jh...@vet.k-state.edu Sent by: histonet-boun...@lists.utsouthwestern.edu 10/29/2010 01:11 PM To histonet histonet@lists.utsouthwestern.edu cc Subject [Histonet] IHC Staining Problem I hope someone can help me with a problem I've been having with my CD79a hand stain. This is a stain that was validated and working perfectly, when a few months ago I started having reduced, to no staining on my controls, with the occasional run staining a little stronger. I followed the discussion not too long ago with the person who was having similar problems, but the suggestions didn't seem to apply (I use Tween 20 in my PBS rinses, and we use APEX slides from Surgipath-not Fischer Plus). I've tried everything I can think of and I hope someone may have some other ideas. For reference this is for a veterinary diagnostic lab, not a human lab, and this is the only stain we've been having this issue with. My protocol is as follows: After deparfinization/rehydration, AR for 20 min in the steamer in Biogenex Citra Solution (I've gotten new Citra Solution, tried a pressure cooker) 5 min peroxide quench, followed by a 10 min protein block using Dako's Protein Block-Serum Free 60 min Primary incubation (Dako CD79a) at 37 C (decreased dilution from 1:100 to 1:65, new bottle of antibody) 30 min incubation at room temp of ImmPRESS anti-Mouse Reagent from Vector Develop stain with DAB chromogen from Vector Washes of PBS/Tween 20 follow between steps Thank you, Jennifer Hill Research Assistant Kansas State University Veterinary Diagnostic Lab ___ 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
