[Histonet] congenital syphilis
Hi, I was hoping someone would have a suggestion for a stain for congenital syphilis on a placenta. Our pathologist wants this done- I know a warthinstarry or steiner would work, but we do not do either of these stains. Would a diff-quik stain work? Thanks, Mandy Bell HTL(ASCP) Community Hospital of the Monterey Peninsula 2 Harris Court Suite B3/B4 Monterey, CA 93940 (831) 647-4791 Confidentiality Notice: This is a transmission from Community Hospital of the Monterey Peninsula. This message and any attached documents may be confidential and contain information protected by state and federal medical privacy statutes. They are intended only for the use of the addressee. If you are not the intended recipient, any disclosure, copying, or distribution of this information is strictly prohibited. If you received this transmission in error, please accept our apologies and notify the sender. Thank you. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Consulting
Ryan Jade, I am inquiring if you may have found a consultant to help you with your dermatology laboratory? Laboratory Concepts, LLC can help you with your needs. I would be happy to speak with you in detail offline. Regards, Amy Farnan a.far...@yahoo.com Disclaimer: The information in this message is confidential. If you are not the intended recipient, do not disclose, copy, or distribute this message, and please immediately contact the sender. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] RE: Re: Methanol in H2O2 explanation
Carl, H202 mixed with Buffer here.(instead of aq.) That's OK too, right? Dana -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Hobbs, Carl Sent: Tuesday, July 10, 2012 11:32 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Re: Methanol in H2O2 explanation Why do some people use methanolic H2O2? Why do people rehydrate after dewaxing? Both are unnecessary, under usual conditions. Methanol was used in the early days as a peroxidase blocker by itself. The combination was devised as a belt and bracer method. As you stated, you use aq H2O2 effectively. So do I and many others. However, for unfixed frozen sections, I would never use aq H2O2, if I wanted sections remaining on my slides After dewaxing, rinse sections in 4x 100% alcohol, rinse in water, endog. Px block while you make up you AR solutions Carl Hobbs Histology and Imaging Manager Wolfson CARD School of Biomedical Sciences Kings College London Guys Campus SE1 1UL Tel: 020 78486813 Fax: 020 78486816 020 78486813 ___ 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] congenital syphilis
Immunohistochemical testing for Treponema pallidum is the preferred method of identification. There are labs (including my own) that offer this. Richard Richard W. Cartun, MS, PhD Director, Histology Immunopathology Director, Biospecimen Collection Programs Assistant Director, Anatomic Pathology Hartford Hospital 80 Seymour Street Hartford, CT 06102 (860) 545-1596 Office (860) 545-2204 Fax Bell, Mandy mandy.b...@chomp.org 7/11/2012 7:31 AM Hi, I was hoping someone would have a suggestion for a stain for congenital syphilis on a placenta. Our pathologist wants this done- I know a warthinstarry or steiner would work, but we do not do either of these stains. Would a diff-quik stain work? Thanks, Mandy Bell HTL(ASCP) Community Hospital of the Monterey Peninsula 2 Harris Court Suite B3/B4 Monterey, CA 93940 (831) 647-4791 Confidentiality Notice: This is a transmission from Community Hospital of the Monterey Peninsula. This message and any attached documents may be confidential and contain information protected by state and federal medical privacy statutes. They are intended only for the use of the addressee. If you are not the intended recipient, any disclosure, copying, or distribution of this information is strictly prohibited. If you received this transmission in error, please accept our apologies and notify the sender. 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
AW: [Histonet] RE: Re: Methanol in H2O2 explanation
For the enzymatic activity of peroxidase it needs an electron-donator (or receptor - I can't find the literature...) in the vicinity; therefore H2O2 and DAB are added ad once, and DAB is oxidized and transformed into the insoluble, amorphe substance through polymerization. Without the donator H2O2 in excess works as inhibitor and blocks the activation-side of the enzyme. I think H2O2 in methanol was primarly preferred, because the frozen slides were fixed at the same time. Rehydration after dewaxing depends on the following reagens. Gudrun -Ursprüngliche Nachricht- Von: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] Im Auftrag von Tony Henwood (SCHN) Gesendet: Mittwoch, 11. Juli 2012 06:21 An: 'Hobbs, Carl'; histonet@lists.utsouthwestern.edu Betreff: [Histonet] RE: Re: Methanol in H2O2 explanation Hi Carl, What do you mean by Why do people rehydrate after dewaxing? Do you really mean that slides do not require rehydration or do you mean that slides can be left to dry after de-waxing prior to staining. Re-hydration is necessary, otherwise xylene will prevent aqueous stains from doing their thing efficiently. I was lead to believe that as H2O2 was catalysed by endogenous peroxidase, the reactive oxygen reacted with the methanol to then degrade the enzyme, but I need to look closer at this chemistry. Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA) Laboratory Manager Senior Scientist Tel: 612 9845 3306 Fax: 612 9845 3318 the children's hospital at westmead Cnr Hawkesbury Road and Hainsworth Street, Westmead Locked Bag 4001, Westmead NSW 2145, AUSTRALIA -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Hobbs, Carl Sent: Wednesday, 11 July 2012 1:32 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Re: Methanol in H2O2 explanation Why do some people use methanolic H2O2? Why do people rehydrate after dewaxing? Both are unnecessary, under usual conditions. Methanol was used in the early days as a peroxidase blocker by itself. The combination was devised as a belt and bracer method. As you stated, you use aq H2O2 effectively. So do I and many others. However, for unfixed frozen sections, I would never use aq H2O2, if I wanted sections remaining on my slides After dewaxing, rinse sections in 4x 100% alcohol, rinse in water, endog. Px block while you make up you AR solutions Carl Hobbs Histology and Imaging Manager Wolfson CARD School of Biomedical Sciences Kings College London Guys Campus SE1 1UL Tel: 020 78486813 Fax: 020 78486816 020 78486813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet * This email and any files transmitted with it are confidential and intended solely for the use of the individual or entity to whom they are addressed. If you are not the intended recipient, please delete it and notify the sender. Views expressed in this message and any attachments are those of the individual sender, and are not necessarily the views of The Children's Hospital at Westmead This note also confirms that this email message has been virus scanned and although no computer viruses were detected, The Childrens Hospital at Westmead accepts no liability for any consequential damage resulting from email containing computer viruses. * ___ 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] Another Microscope Question
Laurie: Köhler illumination is not just observing objects in bright field, it implies the use 2 diaphragms; one near the light source and focused in a way that the diaphragm closest to the light source covers (illuminates) only the area to be observed. With the condenser diaphragm you then increase/decrease the intensity of the light. It is most useful for photomicrography and to obtain the maximum resolution of the numerical aperture (NA) of your objectives. If you look at the slide side wise, you will see that the illuminated area corresponds only to the working filed of each objective. That is why when using Köhler illumination you have to adjust the light source diaphragm every time you change the objective under use. What your pathologists are trying to do is just to increase/reduce the amount of light reaching the object by increasing/reducing the distance of the condenser from the object to be observed. Many years ago and starting in the mid 1920s, Carl Zeiss (and also Ernst Leitz) designed the condensers of their research microscopes with the condenser diaphragm mounted on a slide with a knob. By doing so these condensers could move the condenser diaphragm side wise in a way that the light entered into the condenser not straight from the light source obliquously, and the objects were illuminated side wise. This illumination permitted to observe unstained slides and produced the illusion of three dimension and permitted to observe unstained structures. This type of illumination was quite popular in hematology labs to observe blood smears In 1935 the same Carl Zeiss started to manufacture objectives/condenser sytems using Zernike's principle, and phase microscopy was born. The method of lowering the condenser used by your pathologists has to produce a very poor quality image. You would be better off if you cut a piece of black paper half the diameter of the filter ring of your condenser and place it there. With this opaque filter the light will get to your object side wise, the same as if you have moved the condenser side wise. René J. From: Reilly, Laurie laurie.rei...@jcu.edu.au To: Histonet@lists.utsouthwestern.edu Histonet@lists.utsouthwestern.edu Sent: Tuesday, July 10, 2012 7:21 PM Subject: [Histonet] Another Microscope Question Dear All, My microscope training has been mainly on brightfield microscopes using Koehler illumination. Recently, in discussions with Veterinary Pathologists, they have been advocating winding the condenser way down to the field diaphragm when they are examining urine samples. This goes against the microscopy principles that I have been taught and now teach to our students. Can anyone enlighten me on the value of this practice? Could the same effect be obtained by closing the aperture diaphragm? Thanks in advance for your wisdom. Regards, Laurie. Mr. Laurie REILLY Histopathology School of Veterinary and Biomedical Sciences James Cook University Townsville Qld. 4811 Australia. Phone 07 4781 4468 ___ 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] Re: Methanol in H2O2 explanation
I made a mistakeI DO rehydrate before staining, as Tony pointed out. I just don't use graded alcohols between the xylene and water ( x4 changes 74OP IMS). Carl Hobbs Histology Manager Wolfson Centre for Age-Related Diseases School of Biomedical Sciences King's College London Guys Campus London SE1 1UL Tel.020 7848 6810 fax 020 7848 6816 carl.ho...@kcl.ac.ukmailto:carl.ho...@kcl.ac.uk ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] RE: Re: Methanol in H2O2 explanation
And the reason for that is...??? Beatrice DeBrosse-Serra HT(ASCP)QIHC Isis Pharmaceuticals Antisense Drug Discovery 2855 Gazelle Ct. Carlsbad, CA 92010 760-603-2371 -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Hobbs, Carl Sent: Wednesday, July 11, 2012 7:50 AM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] Re: Methanol in H2O2 explanation I made a mistakeI DO rehydrate before staining, as Tony pointed out. I just don't use graded alcohols between the xylene and water ( x4 changes 74OP IMS). Carl Hobbs Histology Manager Wolfson Centre for Age-Related Diseases School of Biomedical Sciences King's College London Guys Campus London SE1 1UL Tel.020 7848 6810 fax 020 7848 6816 carl.ho...@kcl.ac.ukmailto:carl.ho...@kcl.ac.uk ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
RE: [Histonet] RE: Re: Methanol in H2O2 explanation
It's funny how histology lore gets passed down, passed around and misunderstood over the decades. When I ask questions in labs about why they do a certain procedure they can rarely name a source (besides a particular long-retired particular tech who wrote the original procedure) or give reason as to why it is done that way. Even if it works well, at least people could take the time to understand why! When I was researching buffer compositions long ago I found a dozen variations of PBS alone (and only a dozen because I just stopped looking). When you delve into these things it becomes clear that people made their own solutions for a particular purpose. Maybe they wrote a paper, taught a course or wrote a book. Then that formula was copied by many others and their particular formulation became the state of the art even if the art practiced by someone else had little to do with the original pursuit (the Sheehan book is full of variations of stains that were originally for specific research purposes. There is precious little guidance in any books about what is good for a particular use!). The reason for using methanol / H2O2 that I was told long ago was prevent frozen sections from being damaged by H2O2 bubbling. In fact, we would fix frozens in cold acetone and THEN put the slides in cold methanol/H2O2 (double fixation?) In that lab we never used methanol for deparaffinized sections. Since I had no exposure to the clin lab I never knew about using methanol for blood smear fixation. From literature searches it seems methanol, rather than ethanol, is used primarily because it is cheaper. In other words, it does not appear to be a magical substance! The questions asked here are good ones. However, hopefully those reading and responsible for producing procedure manuals will include the reasoning for each step of the method and not just assume it is necessary, or common knowledge, just because it has been handed down through generations of long suffering techs! Tim Morken -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Gudrun Lang Sent: Wednesday, July 11, 2012 7:17 AM To: histonet@lists.utsouthwestern.edu Subject: AW: [Histonet] RE: Re: Methanol in H2O2 explanation For the enzymatic activity of peroxidase it needs an electron-donator (or receptor - I can't find the literature...) in the vicinity; therefore H2O2 and DAB are added ad once, and DAB is oxidized and transformed into the insoluble, amorphe substance through polymerization. Without the donator H2O2 in excess works as inhibitor and blocks the activation-side of the enzyme. I think H2O2 in methanol was primarly preferred, because the frozen slides were fixed at the same time. Rehydration after dewaxing depends on the following reagens. Gudrun -Ursprüngliche Nachricht- Von: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] Im Auftrag von Tony Henwood (SCHN) Gesendet: Mittwoch, 11. Juli 2012 06:21 An: 'Hobbs, Carl'; histonet@lists.utsouthwestern.edu Betreff: [Histonet] RE: Re: Methanol in H2O2 explanation Hi Carl, What do you mean by Why do people rehydrate after dewaxing? Do you really mean that slides do not require rehydration or do you mean that slides can be left to dry after de-waxing prior to staining. Re-hydration is necessary, otherwise xylene will prevent aqueous stains from doing their thing efficiently. I was lead to believe that as H2O2 was catalysed by endogenous peroxidase, the reactive oxygen reacted with the methanol to then degrade the enzyme, but I need to look closer at this chemistry. Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA) Laboratory Manager Senior Scientist Tel: 612 9845 3306 Fax: 612 9845 3318 the children's hospital at westmead Cnr Hawkesbury Road and Hainsworth Street, Westmead Locked Bag 4001, Westmead NSW 2145, AUSTRALIA -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Hobbs, Carl Sent: Wednesday, 11 July 2012 1:32 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Re: Methanol in H2O2 explanation Why do some people use methanolic H2O2? Why do people rehydrate after dewaxing? Both are unnecessary, under usual conditions. Methanol was used in the early days as a peroxidase blocker by itself. The combination was devised as a belt and bracer method. As you stated, you use aq H2O2 effectively. So do I and many others. However, for unfixed frozen sections, I would never use aq H2O2, if I wanted sections remaining on my slides After dewaxing, rinse sections in 4x 100% alcohol, rinse in water, endog. Px block while you make up you AR solutions Carl Hobbs Histology and Imaging Manager Wolfson CARD School of Biomedical Sciences Kings College London Guys Campus SE1 1UL Tel: 020 78486813 Fax:
[Histonet] Beecher
All ,I know this came up before,but who took over Beecher? We need to order some more punchers. Thanks, Bernice Bernice Frederick HTL (ASCP) Senior Research Tech Pathology Core Facility ECOGPCO-RL Robert. H. Lurie Cancer Center Northwestern University 710 N Fairbanks Court Olson 8-421 Chicago,IL 60611 312-503-3723 b-freder...@northwestern.edumailto:b-freder...@northwestern.edu ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] job opening
We are currently looking for a histotech to join our rapidly growing practice and explore the exciting world of dermatopathology in a new state-of-the-art laboratory overlooking beautiful Lake Michigan. At Dermatology Associates of Wisconsin you will enjoy performing a wide variety of tests including grossing, DIFs, special stains, and IHC. You will experience working 4 days per week for a dermatology group averaging over 30% growth annually for the past 10 years. Benefits include: * 401k match of 100% of the first 4% of employee contribution * Company profit sharing contribution of 7% of employee earnings * Company funded cash balance plan that is a yearly contribution of 2% of employee earnings * Immediate PTO accrual * Leadership that enjoys teaching * A great team atmosphere * Employee discounts * Opportunities for professional growth For more details contact: Human Resources 801 York Street Manitowoc, WI 54220 h...@dermwisconsin.commailto:h...@dermwisconsin.com Office: (920)683-5278 Fax:(920)663-9004 The materials and information in this e-mail are confidential and may contain Protected Health Information covered under the HIPAA Privacy Rule. If you are not the intended recipient, be advised that any unauthorized use, disclosure, copying, distribution, or action taken in reliance on the contents of this information is strictly forbidden by law. If you have received this e-mail in error, please notify me by reply e-mail and then delete this message. Do not pass any of this information to anyone else. Thank you for your cooperation. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] RE: Beecher
http://www.pathologydevices.com/TMArrayer.htm You can contact Ron Gebing at Pathology Devices. Helen -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Bernice Frederick Sent: Wednesday, July 11, 2012 12:14 PM To: Fellow HistoNetters Subject: [Histonet] Beecher All ,I know this came up before,but who took over Beecher? We need to order some more punchers. Thanks, Bernice Bernice Frederick HTL (ASCP) Senior Research Tech Pathology Core Facility ECOGPCO-RL Robert. H. Lurie Cancer Center Northwestern University 710 N Fairbanks Court Olson 8-421 Chicago,IL 60611 312-503-3723 b-freder...@northwestern.edumailto:b-freder...@northwestern.edu ___ 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: Re: Methanol in H2O2 explanation
I agree, lots of lore gets passed down without question or understanding. Many years ago a new PhD who had trained in a good lab came to my lab looking for some NaH. I tried to explain that there was no such chemical when she produced a xeroxed lab protocol calling for NaH to make a malete buffer for EM (that shows you how long ago this was!). So I found the exact recipe but with NaOH. The techs all knew it was a typo but she did not. Geoff On 7/11/2012 12:12 PM, Morken, Timothy wrote: It's funny how histology lore gets passed down, passed around and misunderstood over the decades. When I ask questions in labs about why they do a certain procedure they can rarely name a source (besides a particular long-retired particular tech who wrote the original procedure) or give reason as to why it is done that way. Even if it works well, at least people could take the time to understand why! When I was researching buffer compositions long ago I found a dozen variations of PBS alone (and only a dozen because I just stopped looking). When you delve into these things it becomes clear that people made their own solutions for a particular purpose. Maybe they wrote a paper, taught a course or wrote a book. Then that formula was copied by many others and their particular formulation became the state of the art even if the art practiced by someone else had little to do with the original pursuit (the Sheehan book is full of variations of stains that were originally for specific research purposes. There is precious little guidance in any books about what is good for a particular use!). The reason for using methanol / H2O2 that I was told long ago was prevent frozen sections from being damaged by H2O2 bubbling. In fact, we would fix frozens in cold acetone and THEN put the slides in cold methanol/H2O2 (double fixation?) In that lab we never used methanol for deparaffinized sections. Since I had no exposure to the clin lab I never knew about using methanol for blood smear fixation. From literature searches it seems methanol, rather than ethanol, is used primarily because it is cheaper. In other words, it does not appear to be a magical substance! The questions asked here are good ones. However, hopefully those reading and responsible for producing procedure manuals will include the reasoning for each step of the method and not just assume it is necessary, or common knowledge, just because it has been handed down through generations of long suffering techs! Tim Morken -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Gudrun Lang Sent: Wednesday, July 11, 2012 7:17 AM To: histonet@lists.utsouthwestern.edu Subject: AW: [Histonet] RE: Re: Methanol in H2O2 explanation For the enzymatic activity of peroxidase it needs an electron-donator (or receptor - I can't find the literature...) in the vicinity; therefore H2O2 and DAB are added ad once, and DAB is oxidized and transformed into the insoluble, amorphe substance through polymerization. Without the donator H2O2 in excess works as inhibitor and blocks the activation-side of the enzyme. I think H2O2 in methanol was primarly preferred, because the frozen slides were fixed at the same time. Rehydration after dewaxing depends on the following reagens. Gudrun -Ursprüngliche Nachricht- Von: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] Im Auftrag von Tony Henwood (SCHN) Gesendet: Mittwoch, 11. Juli 2012 06:21 An: 'Hobbs, Carl'; histonet@lists.utsouthwestern.edu Betreff: [Histonet] RE: Re: Methanol in H2O2 explanation Hi Carl, What do you mean by Why do people rehydrate after dewaxing? Do you really mean that slides do not require rehydration or do you mean that slides can be left to dry after de-waxing prior to staining. Re-hydration is necessary, otherwise xylene will prevent aqueous stains from doing their thing efficiently. I was lead to believe that as H2O2 was catalysed by endogenous peroxidase, the reactive oxygen reacted with the methanol to then degrade the enzyme, but I need to look closer at this chemistry. Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA) Laboratory Manager Senior Scientist Tel: 612 9845 3306 Fax: 612 9845 3318 the children's hospital at westmead Cnr Hawkesbury Road and Hainsworth Street, Westmead Locked Bag 4001, Westmead NSW 2145, AUSTRALIA -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Hobbs, Carl Sent: Wednesday, 11 July 2012 1:32 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Re: Methanol in H2O2 explanation Why do some people use methanolic H2O2? Why do people rehydrate after dewaxing? Both are unnecessary, under usual conditions. Methanol was used in the early days as a peroxidase blocker by itself. The
RE: [Histonet] RE: Re: Methanol in H2O2 explanation
Now that's funny! Even if it makes we PhDs look bad. I was once given some tissue in formaldehyde from one of my colleagues in another department to process and stain. The resulting slides looked horrendous and I suspected the fix was the problem. I went back to their lab and asked to see the bottle of formaldehyde which turned out to be formamide. Then I got the shoulder shrug and We thought it was close enough response. Argghh. Brett -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Geoff McAuliffe Sent: Wednesday, July 11, 2012 12:50 PM To: histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] RE: Re: Methanol in H2O2 explanation I agree, lots of lore gets passed down without question or understanding. Many years ago a new PhD who had trained in a good lab came to my lab looking for some NaH. I tried to explain that there was no such chemical when she produced a xeroxed lab protocol calling for NaH to make a malete buffer for EM (that shows you how long ago this was!). So I found the exact recipe but with NaOH. The techs all knew it was a typo but she did not. Geoff On 7/11/2012 12:12 PM, Morken, Timothy wrote: It's funny how histology lore gets passed down, passed around and misunderstood over the decades. When I ask questions in labs about why they do a certain procedure they can rarely name a source (besides a particular long-retired particular tech who wrote the original procedure) or give reason as to why it is done that way. Even if it works well, at least people could take the time to understand why! When I was researching buffer compositions long ago I found a dozen variations of PBS alone (and only a dozen because I just stopped looking). When you delve into these things it becomes clear that people made their own solutions for a particular purpose. Maybe they wrote a paper, taught a course or wrote a book. Then that formula was copied by many others and their particular formulation became the state of the art even if the art practiced by someone else had little to do with the original pursuit (the Sheehan book is full of variations of stains that were originally for specific research purposes. There is precious little guidance in any books about what is good for a particular use!). The reason for using methanol / H2O2 that I was told long ago was prevent frozen sections from being damaged by H2O2 bubbling. In fact, we would fix frozens in cold acetone and THEN put the slides in cold methanol/H2O2 (double fixation?) In that lab we never used methanol for deparaffinized sections. Since I had no exposure to the clin lab I never knew about using methanol for blood smear fixation. From literature searches it seems methanol, rather than ethanol, is used primarily because it is cheaper. In other words, it does not appear to be a magical substance! The questions asked here are good ones. However, hopefully those reading and responsible for producing procedure manuals will include the reasoning for each step of the method and not just assume it is necessary, or common knowledge, just because it has been handed down through generations of long suffering techs! Tim Morken -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Gudrun Lang Sent: Wednesday, July 11, 2012 7:17 AM To: histonet@lists.utsouthwestern.edu Subject: AW: [Histonet] RE: Re: Methanol in H2O2 explanation For the enzymatic activity of peroxidase it needs an electron-donator (or receptor - I can't find the literature...) in the vicinity; therefore H2O2 and DAB are added ad once, and DAB is oxidized and transformed into the insoluble, amorphe substance through polymerization. Without the donator H2O2 in excess works as inhibitor and blocks the activation-side of the enzyme. I think H2O2 in methanol was primarly preferred, because the frozen slides were fixed at the same time. Rehydration after dewaxing depends on the following reagens. Gudrun -Ursprüngliche Nachricht- Von: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] Im Auftrag von Tony Henwood (SCHN) Gesendet: Mittwoch, 11. Juli 2012 06:21 An: 'Hobbs, Carl'; histonet@lists.utsouthwestern.edu Betreff: [Histonet] RE: Re: Methanol in H2O2 explanation Hi Carl, What do you mean by Why do people rehydrate after dewaxing? Do you really mean that slides do not require rehydration or do you mean that slides can be left to dry after de-waxing prior to staining. Re-hydration is necessary, otherwise xylene will prevent aqueous stains from doing their thing efficiently. I was lead to believe that as H2O2 was catalysed by endogenous peroxidase, the reactive oxygen reacted with the methanol to then degrade the enzyme, but I need to look
[Histonet] Antibody Inventory
Good Afternoon Does anyone have any suggestions for primary antibody inventory management? Thank you Mary Helie ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] P63
I guess since Biocare cannot sell their P63 antibody who is carrying it then? Dako, Ventana etc? Karen Heckford HT ASCP CE Lead Histology Technician St. Mary's Medical Center 450 Stanyan St. San Francisco, Ca. 94117 415-668-1000 ext. 6167 karen.heckf...@dignityhealth.org Caution: This email message, including all content and attachments, is CONFIDENTIAL and may be of a nature that is LEGALLY PRIVILEGED. The information contained in this email message is intended only for the use of the recipient(s) named above. If the reader of this message is not the intended recipient or an agent responsible for delivering it to the intended recipient, you have received this document in error. Any further review, dissemination, distribution, or copying of this message is strictly prohibited. If you have received this communication in error, please notify us immediately by reply email. Thank you. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] RE: Beecher
This is where I get my stuff for my TMA machine from http://www.estigen.com/ Loralee McMahon, HTL (ASCP) Immunohistochemistry Supervisor Strong Memorial Hospital Department of Surgical Pathology (585) 275-7210 From: histonet-boun...@lists.utsouthwestern.edu [histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Bernice Frederick [b-freder...@northwestern.edu] Sent: Wednesday, July 11, 2012 12:13 PM To: Fellow HistoNetters Subject: [Histonet] Beecher All ,I know this came up before,but who took over Beecher? We need to order some more punchers. Thanks, Bernice Bernice Frederick HTL (ASCP) Senior Research Tech Pathology Core Facility ECOGPCO-RL Robert. H. Lurie Cancer Center Northwestern University 710 N Fairbanks Court Olson 8-421 Chicago,IL 60611 312-503-3723 b-freder...@northwestern.edumailto:b-freder...@northwestern.edu ___ 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] Re: Methanol in H2O2 explanation
Ha! I've found it. It can be found in Dr. Kiernan's book in the chapter enzymehistochemistry - the thing, that an electron-donator is used for demonstration. And he states, that methanol alone also inhibits peroxidase - perhaps by the fixation = denaturation effect on the protein? In the following publication one can see the equations for the peroxidase-reaction and this phrase: In this study, we have used a steady state kinetics approach to demonstrate that MPO is irreversibly inactivated by excess H2O2 in the absence of reducing substrate. The Open Enzyme Inhibition Journal, 2009, 2, 28-35 Mechanism-Based Inhibition of Myeloperoxidase by Hydrogen Peroxide: Enhancement of Inactivation Rate by Organic Donor Substrates It's rather exhausting to prove the own statements ;-) puh. Gudrun -Ursprüngliche Nachricht- Von: Morken, Timothy [mailto:timothy.mor...@ucsfmedctr.org] Gesendet: Mittwoch, 11. Juli 2012 18:12 An: gu.l...@gmx.at; histonet@lists.utsouthwestern.edu Betreff: RE: [Histonet] RE: Re: Methanol in H2O2 explanation It's funny how histology lore gets passed down, passed around and misunderstood over the decades. When I ask questions in labs about why they do a certain procedure they can rarely name a source (besides a particular long-retired particular tech who wrote the original procedure) or give reason as to why it is done that way. Even if it works well, at least people could take the time to understand why! When I was researching buffer compositions long ago I found a dozen variations of PBS alone (and only a dozen because I just stopped looking). When you delve into these things it becomes clear that people made their own solutions for a particular purpose. Maybe they wrote a paper, taught a course or wrote a book. Then that formula was copied by many others and their particular formulation became the state of the art even if the art practiced by someone else had little to do with the original pursuit (the Sheehan book is full of variations of stains that were originally for specific research purposes. There is precious little guidance in any books about what is good for a particular use!). The reason for using methanol / H2O2 that I was told long ago was prevent frozen sections from being damaged by H2O2 bubbling. In fact, we would fix frozens in cold acetone and THEN put the slides in cold methanol/H2O2 (double fixation?) In that lab we never used methanol for deparaffinized sections. Since I had no exposure to the clin lab I never knew about using methanol for blood smear fixation. From literature searches it seems methanol, rather than ethanol, is used primarily because it is cheaper. In other words, it does not appear to be a magical substance! The questions asked here are good ones. However, hopefully those reading and responsible for producing procedure manuals will include the reasoning for each step of the method and not just assume it is necessary, or common knowledge, just because it has been handed down through generations of long suffering techs! Tim Morken -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Gudrun Lang Sent: Wednesday, July 11, 2012 7:17 AM To: histonet@lists.utsouthwestern.edu Subject: AW: [Histonet] RE: Re: Methanol in H2O2 explanation For the enzymatic activity of peroxidase it needs an electron-donator (or receptor - I can't find the literature...) in the vicinity; therefore H2O2 and DAB are added ad once, and DAB is oxidized and transformed into the insoluble, amorphe substance through polymerization. Without the donator H2O2 in excess works as inhibitor and blocks the activation-side of the enzyme. I think H2O2 in methanol was primarly preferred, because the frozen slides were fixed at the same time. Rehydration after dewaxing depends on the following reagens. Gudrun -Ursprüngliche Nachricht- Von: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] Im Auftrag von Tony Henwood (SCHN) Gesendet: Mittwoch, 11. Juli 2012 06:21 An: 'Hobbs, Carl'; histonet@lists.utsouthwestern.edu Betreff: [Histonet] RE: Re: Methanol in H2O2 explanation Hi Carl, What do you mean by Why do people rehydrate after dewaxing? Do you really mean that slides do not require rehydration or do you mean that slides can be left to dry after de-waxing prior to staining. Re-hydration is necessary, otherwise xylene will prevent aqueous stains from doing their thing efficiently. I was lead to believe that as H2O2 was catalysed by endogenous peroxidase, the reactive oxygen reacted with the methanol to then degrade the enzyme, but I need to look closer at this chemistry. Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA) Laboratory Manager Senior Scientist Tel: 612 9845 3306 Fax: 612 9845 3318 the children's hospital at westmead Cnr Hawkesbury Road and Hainsworth Street, Westmead Locked Bag 4001, Westmead
[Histonet] RE: Antibody Inventory
We made up an excel spread sheet that contains: Antibody/Ancillary solutions(buffer, detection kits) Company Source Catalog # Lot # Expiration Date When we open an new antibody, we clear out the Lot # Expiration date and add it to the re-order list, ensuring that we never run out. Then when the new antibody comes in, we can quickly add it to the supply list. Additionally, the techs in the department found that it helps them to assign a color to each expiration month. So the expiration date cell will be; blue for January, red for Februaryand so on to give a visual help when watching for upcoming expiring antibodies. They also made a cheat sheet of colors that correspond to each month. I have also found that this list helps me immensly when inventory comes around!! Always did hate fishing around in the 'frig searching and counting antibodies!! I'm lucky, my department is very good about keeping up with the incoming/outgoing stock. hope this helps! :) Lynette Lynette Pavelich, HT(ASCP) Histology Supervisor Hurley Medical Center One Hurley Plaza Flint, MI 48503 ph: 810.262.9948 mobile: 810.444.7966 From: histonet-boun...@lists.utsouthwestern.edu [histonet-boun...@lists.utsouthwestern.edu] on behalf of Helie, Mary [mary.he...@yale.edu] Sent: Wednesday, July 11, 2012 1:36 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Antibody Inventory Good Afternoon Does anyone have any suggestions for primary antibody inventory management? Thank you Mary Helie ___ 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] RE: P63
They are still selling it. But they do not know for how long. And because it is a legal issue they can't tell me whom is going to be the sole provider either. Loralee McMahon, HTL (ASCP) Immunohistochemistry Supervisor Strong Memorial Hospital Department of Surgical Pathology (585) 275-7210 From: histonet-boun...@lists.utsouthwestern.edu [histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Heckford, Karen - SMMC-SF [karen.heckf...@dignityhealth.org] Sent: Wednesday, July 11, 2012 1:37 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] P63 I guess since Biocare cannot sell their P63 antibody who is carrying it then? Dako, Ventana etc? Karen Heckford HT ASCP CE Lead Histology Technician St. Mary's Medical Center 450 Stanyan St. San Francisco, Ca. 94117 415-668-1000 ext. 6167 karen.heckf...@dignityhealth.org Caution: This email message, including all content and attachments, is CONFIDENTIAL and may be of a nature that is LEGALLY PRIVILEGED. The information contained in this email message is intended only for the use of the recipient(s) named above. If the reader of this message is not the intended recipient or an agent responsible for delivering it to the intended recipient, you have received this document in error. Any further review, dissemination, distribution, or copying of this message is strictly prohibited. If you have received this communication in error, please notify us immediately by reply email. 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] Methanol in H2O2 explanation--Summary
Hi Histonetters, This has been a fun conversation. I called up a technical support scientist at Vector Labs with the question and was told basically let me get back to you on that. :) But with some digging and the reference from Gudrun (thanks!) I've pieced together the following narrative. 1) Excess H2O2 leads to accumulation of Peroxidase compound III. The Third Intermediate Compound of Horseradish Peroxidase and Hydrogen Peroxide Philip George J. Biol. Chem. 1953 201: 427-434. www.jbc.org/content/201/1/427.full.pdf 2) Compound III is inactive as a peroxidase and can transfer of electron to hydrogen peroxide forming hydroxyl radical which attacks the heme porphoryin ring causing irreversible inactivation of the enzyme. Femji et-al The Open Enzyme Inhibition Journal, 2009, 2, 28-35 3) Methanol itself effectively (ethanol not so much) cleaves Heme groups from Horseradish peroxidase. Werner Straus J Histochem Cytochem, 1974 22:908 4) As the Vector Labs Quenching Protocols cheat sheet explains: Methanol accelerates the destruction of the heme groups so a lower concentration of H2O2 can be used for a longer period of time. www.vectorlabs.com/Protocols/Supprotocols/quenchHRP.pdf So Methanol actually is a magic chemical in this case! Jerry Ricks Research Scientist University of Washington Department of Pathology From: gu.l...@gmx.at To: histonet@lists.utsouthwestern.edu Date: Wed, 11 Jul 2012 20:22:38 +0200 Subject: [Histonet] Re: Methanol in H2O2 explanation Ha! I've found it. It can be found in Dr. Kiernan's book in the chapter enzymehistochemistry - the thing, that an electron-donator is used for demonstration. And he states, that methanol alone also inhibits peroxidase - perhaps by the fixation = denaturation effect on the protein? In the following publication one can see the equations for the peroxidase-reaction and this phrase: In this study, we have used a steady state kinetics approach to demonstrate that MPO is irreversibly inactivated by excess H2O2 in the absence of reducing substrate. The Open Enzyme Inhibition Journal, 2009, 2, 28-35 Mechanism-Based Inhibition of Myeloperoxidase by Hydrogen Peroxide: Enhancement of Inactivation Rate by Organic Donor Substrates It's rather exhausting to prove the own statements ;-) puh. Gudrun -Ursprüngliche Nachricht- Von: Morken, Timothy [mailto:timothy.mor...@ucsfmedctr.org] Gesendet: Mittwoch, 11. Juli 2012 18:12 An: gu.l...@gmx.at; histonet@lists.utsouthwestern.edu Betreff: RE: [Histonet] RE: Re: Methanol in H2O2 explanation It's funny how histology lore gets passed down, passed around and misunderstood over the decades. When I ask questions in labs about why they do a certain procedure they can rarely name a source (besides a particular long-retired particular tech who wrote the original procedure) or give reason as to why it is done that way. Even if it works well, at least people could take the time to understand why! When I was researching buffer compositions long ago I found a dozen variations of PBS alone (and only a dozen because I just stopped looking). When you delve into these things it becomes clear that people made their own solutions for a particular purpose. Maybe they wrote a paper, taught a course or wrote a book. Then that formula was copied by many others and their particular formulation became the state of the art even if the art practiced by someone else had little to do with the original pursuit (the Sheehan book is full of variations of stains that were originally for specific research purposes. There is precious little guidance in any books about what is good for a particular use!). The reason for using methanol / H2O2 that I was told long ago was prevent frozen sections from being damaged by H2O2 bubbling. In fact, we would fix frozens in cold acetone and THEN put the slides in cold methanol/H2O2 (double fixation?) In that lab we never used methanol for deparaffinized sections. Since I had no exposure to the clin lab I never knew about using methanol for blood smear fixation. From literature searches it seems methanol, rather than ethanol, is used primarily because it is cheaper. In other words, it does not appear to be a magical substance! The questions asked here are good ones. However, hopefully those reading and responsible for producing procedure manuals will include the reasoning for each step of the method and not just assume it is necessary, or common knowledge, just because it has been handed down through generations of long suffering techs! Tim Morken -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Gudrun Lang Sent: Wednesday, July 11, 2012 7:17 AM To: histonet@lists.utsouthwestern.edu Subject: AW: [Histonet] RE: Re: Methanol in H2O2 explanation For the
[Histonet] Mohs Histotech Opening- 4 Day Work Week- Sarasota, FL
Allied Search Partners is working with a laboratory in the area of Sarasota, FL looking for a qualified and licensed Histotechnologist or Histotechnician with at least one year of experience with Mohs. Also available in the state of FL- a Histology Manager position. Visit our Careers page for details! http://www.alliedsearchpartners.com/careers.php To Apply Please email or fax resume to bran...@alliedsearchpartners.com mailto:bran...@alliedsearchpartners.com or fax to 888 388 7572 tel:888%20388%207572 . No other information is given about location or the organization at this time. Please send resume for review by our recruiters and all qualified candidates will be submitted to HR for further review. Once we have had time to further consider your application, then more information will be made available. Thank you! Position: Histotechnician or Histotechnologist Schedule: Full time/4 day work week Location: Sarasota, FL area Excellent pay and benefits! Requirements: At least 1 year working experience with Mohs and processing of permanent sections HT/HTL certification by ASCP FL Clinical Lab License -- Brannon Owens Recruitment Manager Allied Search Partners T: 888.388.7571 ext. 106 F: 888.388.7572 To view a complete list of Allied Search Partners current openings go to: http://www.alliedsearchpartners.com/careers.php LinkedIn: http://www.linkedin.com/pub/brannon-owens/28/528/823 *If you wish to no longer receive emails from Allied Search Partners please respond to this email message with remove. This email including its attachments is intended only for the confidential use of the individual to whom it is addressed. If you are not the intended recipient, any use, dissemination, distribution or copying of this message or its attachments is prohibited. If you have received this message in error, please notify us immediately, and delete this message and its attachments permanently from your system. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Diane Tokugawa/CA/KAIPERM is out of the office.
I will be out of the office starting 07/11/2012 and will not return until 07/18/2012. Note: For Cytology issues, please call Molly at 8-421-5487 or Wanda 8-421-5426 For Histology issues, please call Barbara at 8-421-4959, IHC/Histo issues Kiran at 8-421-5404, or general histology client service at 8-421-5408. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Thank you for CD40 suggestions!
Thank you everybody who gave me excellent suggestions and feedback on CD40 on frozen sections, especially Gayle! I will keep the individuals posted on how everything goes. Thanks, Bea Beatrice DeBrosse-Serra HT(ASCP)QIHC Isis Pharmaceuticals Antisense Drug Discovery 2855 Gazelle Ct. Carlsbad, CA 92010 760-603-2371 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] (no subject)
http://linkgolfscore.com/wp-admin/likeit.php?unit72.html ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
RE: [Histonet] Re: Methanol in H2O2 explanation
So, the thinking at the time was to kill the endogenous peroxidase twice! No one can say that us oldies weren't thorough! Regards Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA) Laboratory Manager Senior Scientist Tel: 612 9845 3306 Fax: 612 9845 3318 the children's hospital at westmead Cnr Hawkesbury Road and Hainsworth Street, Westmead Locked Bag 4001, Westmead NSW 2145, AUSTRALIA -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Gudrun Lang Sent: Thursday, 12 July 2012 4:23 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Re: Methanol in H2O2 explanation Ha! I've found it. It can be found in Dr. Kiernan's book in the chapter enzymehistochemistry - the thing, that an electron-donator is used for demonstration. And he states, that methanol alone also inhibits peroxidase - perhaps by the fixation = denaturation effect on the protein? In the following publication one can see the equations for the peroxidase-reaction and this phrase: In this study, we have used a steady state kinetics approach to demonstrate that MPO is irreversibly inactivated by excess H2O2 in the absence of reducing substrate. The Open Enzyme Inhibition Journal, 2009, 2, 28-35 Mechanism-Based Inhibition of Myeloperoxidase by Hydrogen Peroxide: Enhancement of Inactivation Rate by Organic Donor Substrates It's rather exhausting to prove the own statements ;-) puh. Gudrun -Ursprüngliche Nachricht- Von: Morken, Timothy [mailto:timothy.mor...@ucsfmedctr.org] Gesendet: Mittwoch, 11. Juli 2012 18:12 An: gu.l...@gmx.at; histonet@lists.utsouthwestern.edu Betreff: RE: [Histonet] RE: Re: Methanol in H2O2 explanation It's funny how histology lore gets passed down, passed around and misunderstood over the decades. When I ask questions in labs about why they do a certain procedure they can rarely name a source (besides a particular long-retired particular tech who wrote the original procedure) or give reason as to why it is done that way. Even if it works well, at least people could take the time to understand why! When I was researching buffer compositions long ago I found a dozen variations of PBS alone (and only a dozen because I just stopped looking). When you delve into these things it becomes clear that people made their own solutions for a particular purpose. Maybe they wrote a paper, taught a course or wrote a book. Then that formula was copied by many others and their particular formulation became the state of the art even if the art practiced by someone else had little to do with the original pursuit (the Sheehan book is full of variations of stains that were originally for specific research purposes. There is precious little guidance in any books about what is good for a particular use!). The reason for using methanol / H2O2 that I was told long ago was prevent frozen sections from being damaged by H2O2 bubbling. In fact, we would fix frozens in cold acetone and THEN put the slides in cold methanol/H2O2 (double fixation?) In that lab we never used methanol for deparaffinized sections. Since I had no exposure to the clin lab I never knew about using methanol for blood smear fixation. From literature searches it seems methanol, rather than ethanol, is used primarily because it is cheaper. In other words, it does not appear to be a magical substance! The questions asked here are good ones. However, hopefully those reading and responsible for producing procedure manuals will include the reasoning for each step of the method and not just assume it is necessary, or common knowledge, just because it has been handed down through generations of long suffering techs! Tim Morken -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Gudrun Lang Sent: Wednesday, July 11, 2012 7:17 AM To: histonet@lists.utsouthwestern.edu Subject: AW: [Histonet] RE: Re: Methanol in H2O2 explanation For the enzymatic activity of peroxidase it needs an electron-donator (or receptor - I can't find the literature...) in the vicinity; therefore H2O2 and DAB are added ad once, and DAB is oxidized and transformed into the insoluble, amorphe substance through polymerization. Without the donator H2O2 in excess works as inhibitor and blocks the activation-side of the enzyme. I think H2O2 in methanol was primarly preferred, because the frozen slides were fixed at the same time. Rehydration after dewaxing depends on the following reagens. Gudrun -Ursprüngliche Nachricht- Von: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] Im Auftrag von Tony Henwood (SCHN) Gesendet: Mittwoch, 11. Juli 2012 06:21 An: 'Hobbs, Carl'; histonet@lists.utsouthwestern.edu Betreff: [Histonet] RE: Re: Methanol in H2O2 explanation Hi Carl, What do you mean by Why do people
