Re: [Histonet] [EXTERNAL] Epredia xylene substitute

Hi Ann, we have used ShellSol for years in our VIPs until our supplier stopped it. That's also a kind of naphta with aliphatic hydrogencarbons. After that wie changed to the Sakura-substitute with the same protocol. For standard processing we had/have two stations with 60 min each at 40 °C. For

Re: [Histonet] Tissue Processor Down Question

Depending on the time the cassettes were in clearing medium, I would transfer them into molten paraffin at least for double of the time in the processor. And then embed etc. Try a few blocks to see, if infiltration was sufficient. If cutting is hampered, prolong the time in paraffin. Gudrun

Re: [Histonet] preordering giemsa stains

In my place it is ordered on demand, but the pathologists usually perfer IHC for Hp. Gudrun -Ursprüngliche Nachricht- Von: Paula via Histonet [mailto:histonet@lists.utsouthwestern.edu] Gesendet: Dienstag, 27. Februar 2024 20:46 An: histonet@lists.utsouthwestern.edu Betreff: [Histonet]

[Histonet] microwave processing

Dear histonetters! I have a question for those who have an insight in the whole landscape of pathologies. I am reading the microwave application book of Dr. Leong (2009). He writes very enthusiastic about fixation and processing with mircrowaves. I know there are microwave processors for

Re: [Histonet] tissue cassettes

Hi, Since we have turned to embedding centers in the late 80ies we let the cassettes sit in the centers without additional paraffin. We only see such "jumping out" tissue, when the cassettes are not warmed (let the lid open) and the tissue renders too cold. As a result tissue and paraffin don't

Re: [Histonet] Bone samples

Hi, In my opinion the hardness of the decalcified blocks is often rather due to the paraffin-processing than the residual calcium. Especially when the tissue is decalcified really long. The hardness comes from the dehydration and "cooking" of collagen fibers. So additional decal will not help

Re: [Histonet] Detergent in heating antigen retrieval

Hi, I think the answer is as so often: it depends. Detergens solves membranes partly and leads to a higher permeabilization of the tissue. Some antigens may take advantage of that, some may not need it. Higher permeability is good for detection with high molecular complexes. Gudrun Lang

Re: [Histonet] Prostate biopsy-water logged popping /lost at cutting?

What first comes in my mind is, that the tissue was a little too cold when placed into the embedding mold. That would cause an insufficient junction with the paraffin. If there was not enough hot paraffin in the embedding mold and handling took a little bit too long, or if the tissue was rather

[Histonet] shelf life of working antibody solutions in IHC

Hi all! I have tried to find a general instruction for the shelf life of antibody working solutions. With automated IHC you usually fill the container with the working solution and depending on the frequency of usage they stay on the instrument at roomtemperature (or higher) or are put in the

[Histonet] history of H staining

Hi all! Does anybody know, when the H stain became that dominant routine-stain in the pathology labs? It was introduced by Wissowzky 1876, but I am curious when our usual histoprocess became worldwide standard. Regards Gudrun Lang ___ Histonet

[Histonet] cutting with glassknifes

Dear histonetters! I have a question for the experienced histo-people. Is it still in practice to use glassknifes for cutting on rotary- or sliding-microtomes? For plastic-embedded tissue in light-microscopy? Or ar glassknifes only found in EM-technique? I am just curious, because I assume,

Re: [Histonet] Sudanblack B on FFPET

Many thanks to all for your helpful hints. Kind regards Gudrun Lang -Ursprüngliche Nachricht- Von: Tony Henwood via Histonet [mailto:histonet@lists.utsouthwestern.edu] Gesendet: Dienstag, 28. März 2023 21:13 An: Bob Richmond; Histonet@lists.utsouthwestern.edu Betreff: Re: [Histonet]

[Histonet] Sudanblack B on FFPET

Hallo! Has anyone experience with Sudanblack B on paraffin slides for staining lipofuszin? A doctor wants the demonstration of the lipoid content of foamy cells or granulocytes in lung. I've found protocols that have incubation-times from 10 minutes to over-night. I've made a saturated

Re: [Histonet] destain for IHC

We do restaining for IHC also, but without destaining at all. Hematoxylin doesn't matter because nuclei are stained after IHC with hematoxylin anyway. And Eosin gets lost through all the washing in buffer. The problem is, that HE section are usually mounted on non-adhesive slides, and they float

[Histonet] Alcianblue for fast frozen sections

Dear histonetters! Today I've heard about alcianblue staining on fast frozen sections, but I've got no details. I would like to know, if the staining result is the same as for staining AB on paraffinslides. They use the stain on transplantation liver. Is this a usual procedure? I would be

Re: [Histonet] Reconstituting antibodies and dilutions

500 µg in 1000µl is the same as 100µg in 200µl So I would reconstitute the 100 µg with 200 µl of water to get the liquid antibody-concentrate. Then I would make the working-dilution: eg. 5µl of concentrate + 5000µl buffer The correct way would be 5 µl + 4995 µl, but in practice this does'nt

Re: [Histonet] Absence of Crystals in stains

My first thought about calciumcrystals is solving by the chromatic acid. I would recommend a vonKossa stain for calcium deposits. Regards Gudrun Lang -Ursprüngliche Nachricht- Von: Akemi via Histonet [mailto:histonet@lists.utsouthwestern.edu] Gesendet: Dienstag, 9. August 2022 23:51

[Histonet] Looking for old Microm-bladeholders

Dear histonetters, I am looking for old Microm bladeholders for sliding-microtomes. Those ones, that have a shorter coverplate than the standard blade. We prefer them for working, but they aren't sold any more. I would be happy for any idea or contact - preferable in Europe. Maybe someone has an

Re: [Histonet] Prussian Blue Reaction

Hi Jennifer, Why do you want to reduce the staining? I ask, because the impact of hydrochloric acid on the tissue may influence the following results anyway. I think, that the prussian blue pigment cannot be removed in an easy way. It is resistent to solvents and mineral acids.

Re: [Histonet] hexamethyleneimine

Jones Methenamin Silver Impregnation, for staining basalmembrans in glomeruli. Gudrun -Ursprüngliche Nachricht- Von: LEROY H BROWN via Histonet [mailto:histonet@lists.utsouthwestern.edu] Gesendet: Montag, 12. Oktober 2020 00:45 An: histonet@lists.utsouthwestern.edu Betreff: Re:

Re: [Histonet] Spectra

Hi! Do you use your own reagens or the Leica Spectra dyes? We have house-made ones. Too faint eosin staining can be rendered more intense with a few drops of acetic acid into the eosin to reach a pH about 5. Second, it is often a prolonged washing after the eosin that causes excess stain to be

Re: [Histonet] Question concerning H. pylori staining

Hi! I found this instruction for a Hp- stain, that sounds similiar to yours. They want the slides to be airdried after water-rinsing and before xylen. But the result should be blue bacteria, not purple. I would try to let the slides air-dry for about half a minute, then rinse very-very short in

[Histonet] FISH on BondMax

Dear all! I would appreciate input on the FISH-function of the BondMax from Leica. Is this feature reliable and better/as good as manual staining? Would anyone recommend this instrument for FISH staining? Thank you Lang Gudrun ___ Histonet

Re: [Histonet] Stainer vs. Stainer

Hi Terri, Have you actually used the Spectra from Leica? This is the follower of ST5020 and CV5020. I am also interested in some feedback on this instrument. Gudrun Lang -Ursprüngliche Nachricht- Von: Terri Braud via Histonet [mailto:histonet@lists.utsouthwestern.edu] Gesendet:

Re: [Histonet] FISH question

e you have digested long enough with pepsin? If the tissue is not well digested you will see background. We use sodium thiocyanate for pretreatment reagent, not citric buffer. These are my first thoughts. Mark On Tue, Feb 19, 2019 at 9:18 AM Gudrun Lang via Histonet < histonet@lists.utsouthweste

[Histonet] FISH question

Dear histonetters! I have difficulties with my FISH preparation on FFPET. I struggle with massive background. It looks like a thick fluorescent film. The signals can't be seen because of the background. Even the nuclei are hard to see. The background is within the tissue but also surrounds it.

Re: [Histonet] VIP issue

are forced to end using ShellSol, because the company stops the production. We don't like to switch to xylene. Which xylen-substitutes are recommended for the VIP besides of Tissue-Clear (Sakura)? Best wishes Gudrun -Ursprüngliche Nachricht- Von: Gudrun Lang via Histonet [mailto:histonet

[Histonet] VIP issue

Dear all! I have a question for those, who are familiar with the VIPs from Sakura. Last time we changed the reagenses the cleaning-ethanol (96%) was very milky and even was full of many small particles. It was a paraffin-soup. What is the cause for such a case? We have been using the organic

Re: [Histonet] Quality Measure for Pathology

Linda, that's exactly what we do here. I wondered, if you fixed a cut-off. Because in quality-audits they often asked the "what do you if.."-questions. for example: "There is an increase in the number of discrepancies in this special month. The rate is over your defined aim. What did you do to

Re: [Histonet] Quality Measure for Pathology

Linda, How do you handle this key figure of discrepancies? Have you set a cut-off (per person, per day) for any consequencies? What are the consequencies, besides repeated training? thanks Gudrun -Ursprüngliche Nachricht- Von: Blazek, Linda via Histonet

[Histonet] inhouse procedures in histotechnics

Hi! What would you say is an inhouse-procedure in a typical clinical histo-lab. The question is regarded to accredidation in quality management. I'm asked to make a list of all inhouse-procedures. Special staining protocols? Immunhisto on stainer? I think histolabs are hardly comparable

Re: [Histonet] Bouin postfixation

The treatment with bouin afterwards doesn't make the formaldehyde-fixation undone. This would be rather a kind of antigen-retrieval and results at least in a mixture of both fixations. I would stay with the familiar formaldehyde-fixation without experiments and try to get the best out of it

Re: [Histonet] RET IHC testing

I bought it from Cellsignalling (monoclonal rabbit), but just started with the procedure on lung tissue with no experience until now. I took ileum as positiv control, with stained Peyer-plaques as result. Gudrun Lang -Ursprüngliche Nachricht- Von: Charles Riley via Histonet

Re: [Histonet] V600E

This is an mutationspecific antibody against BRAF. And the mutation detected is called p.V600E. The exchange of an nucleotid in the DNA (codon 600) leads to an tranformed and always activated BRAF-protein, that is sensitive to an thyrosinkinase-inhibitor as an eg. Melanoma therapy. As far as I

Re: [Histonet] Masson's Trichrome Troubleshooting

Hi, Check the staining after the PTA/PMA step. The acid should destain all the tissue areas, where afterwards the anilinblue should bind. The acid replaces the Biebrich scarlett in this areas and enables and enhances the binding of anilinblue. If the differentiation in insufficient, it may be the

[Histonet] new developments in histopathology?

Dear histonetters, I have to do financial planning for our diagnostic histolab until 2023 for the necessitiy of new instruments etc. As far as diagnostic development is a rather fast thing, I hardly can imagine what will aproach us in 5 years in the clinical set. What do you think? Which

Re: [Histonet] Modified GMS Protocols

Biological stains were mainly found by trial and error through the century. I think the main goals were to find a good result (for the purpose) within the available resources (cheap / expensive, easy/difficult to get) and with a practical application. Later also health-concerns played a role,

Re: [Histonet] gross photography

Hi, We use MakroPath from Milestone in a routine histolab. The camera is mounted on the top oft he grossing-station, with an integrated PC+monitor and pedals for zooming and taking photos. Within this system you can mark the pictures, draw something, measure something ... In comparison to the

[Histonet] Ventana special stainer

Hello! I have a question for those, who have experience with the "old" Ventana special stainer Nexes and switched to the "new" special stainer Benchmark. Is there a difference in the quality of the stainings? Better reagenses? More possibilities to adapt the protocols? Or is it just the same and

[Histonet] decal protocol with EDTA for bonemarrow biopsies

Dear histonetters! I would be happy about some input about decalcification protocols with EDTA of trephine bonemarrow biopsies. recommended duration of fixation? recommended duration of decalcification? strategies for speed-up of decal? recommended EDTA-solution formula? Hopefully some

Re: [Histonet] Digital Pathology & Coverslipping

Hi! We scan our daily HEs for 1,5 year now. We have a Leica glass-coverslipper. The brilliance of the images is very good. With film we had the experience, that the brilliance of digital photos were never of the same quality as with glass. (and had always issues with coming off the film after a

Re: [Histonet] Tissue Fixation

I second the opinion of Joyce. We see such effects in portio-conisations, that are done with a thermo-electrical knife. The surface and the underlying area show a very pink colour in HE. It can also be seen in prostata-chips. IHC on such biopsies shows the effect of an non-stainable edge with a

[Histonet] WG: question about retina IHC

Because I have no experience with this special specimens, I give the question below to the kind histonet and its professionals. Gudrun Lang Von: Amirkavei Mooud [mailto:mooud.amirka...@aalto.fi] Gesendet: Donnerstag, 23. März 2017 13:02 An: Lang Gudrun Betreff: question about retina IHC

Re: [Histonet] staining samples years in formalin

Hi Mary Lou, I think the problem is, that proteoglycanes will be solved by acids through decal. Try EDTA decal or prolong the staining time in Alcianblue - maybe for 2 hours? Try to decal your controls in the same manner and compare the results. I don't think, that formalin fixation is the

Re: [Histonet] Questions RE: "what I have found about p16" and cellblocks for controls

Your concerns are reasonable. Cyto-specimens are usually fixed in alcoholic solutions not in NBF. Alcohol-fixation gives false positives in Her2. This can also be seen in underfixed tissue, that is mainly fixed by the ethanols in the processor. Her2-protein is a normal protein on the cellsurface.

Re: [Histonet] Staining stomach cells

Did you look at the mucin-antibodies? Pepsinogen-IHC should be positiv in chief-cells. Gastrin-IHC should be positiv in parietal-cells. Chromogranin A should be positiv in endocrine cells. just my ideas. please confirm with your own research. Gudrun -Ursprüngliche Nachricht- Von: Judi

Re: [Histonet] PAS/Decal Question

Hi Pam, my personal opinion is, that 2 hours fixation is too short for sufficient tissue-protection before acid decalcification. Formic acid at 50°C must have an impact on glycoproteins. Wether it is a kind of solving the "sugars" or beginning oxidation of the OH-groups (like periodic acid does in

Re: [Histonet] PAS Stain

As far as I remember the incubation temperature is at roomtemperature. 60°C would rather denature the native enzyme than increase activity. Look at the optimal working temp of the reagens you have bought. regards Gudrun -Ursprüngliche Nachricht- Von: Joanne Clark via Histonet

[Histonet] WG: copper stain

Thank you all for your responses. Gudrun -Ursprüngliche Nachricht- Von: Gudrun Lang via Histonet [mailto:histonet@lists.utsouthwestern.edu] Gesendet: Mittwoch, 20. April 2016 18:42 An: histonet@lists.utsouthwestern.edu Betreff: [Histonet] copper stain Hi all! Which stain would you

[Histonet] copper stain

Hi all! Which stain would you prefer to demonstrate copper? Rhodanin or Victoria blue? Thanks in advance Gudrun ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet

Re: [Histonet] formalin and shrinkage

to keep things flat and oriented. Some people don't like them because of carryover. I just say change your processor reagents more often. Sincerely, Jay A. Lundgren, M.S., HTL (ASCP) On Mon, Feb 22, 2016 at 9:59 AM, Gudrun Lang via Histonet <histonet@lists.utsouthwestern.

Re: [Histonet] Davidson's fixative and IHC question

I think fixing with Davidson is ethanol-fixation in the first line. There are some antigens, that are susceptible to ethanol. Gudrun -Ursprüngliche Nachricht- Von: Judi Ford via Histonet [mailto:histonet@lists.utsouthwestern.edu] Gesendet: Montag, 22. Februar 2016 18:45 An:

[Histonet] formalin and shrinkage

Hi! Today someone asked me about shrinkage caused by the fixation with formaldehyde specially on skin-biopsies. She spoke about shrinkage of 30% percent. In my opinion shrinkage is mainly caused by the processing with dehydration and defatting. Formaldehyde renders the tissue harder but not

Re: [Histonet] Double stain IHC question

In my opinion, this would only be possible, if the commercial and the homegrown antibody are from different species. For example one from mouse and one from rabbit. Then you can proceed with different secondaries (goat anti mouse conjugated with peroxidase, goat anti rabbit conjugated with

Re: [Histonet] keratohyalin granules

Do you stain with hematoxylin only? or also with eosin. I think eosin is the dye, that would highlight the basic proteins in the granula. Gudrun -Ursprüngliche Nachricht- Von: Kalleberg, Kristopher via Histonet [mailto:histonet@lists.utsouthwestern.edu] Gesendet: Mittwoch, 17. Februar

Re: [Histonet] Sponges for processing biopsies

Sponges are available in different qualities. We use very soft ones, that don't hurt the tissue. I know, there are also very hard materials on the market, that may render holes in the underfixed biopsies. Gudrun -Ursprüngliche Nachricht- Von: Lester Raff MD via Histonet

Re: [Histonet] Nuclear bubbling artifact

I've read about a group, that observed living cells during the fixation-process. They saw bubbling in the first period of contact and penetration of formaldehyde. After a certain time the bubbles disappeared again. Along this observation for me bubbles are a sign of too short fixation. Gudrun

[Histonet] additional barcode on ventana-labels

Hi! Did anyone manage to bring Ventana to open their label-design? We need an additional 2D-Barcode with the slide-data on the label. Regards Gudrun Lang ___ Histonet mailing list Histonet@lists.utsouthwestern.edu

[Histonet] Hirschsprung diagnostic kit

Hi all! Has anyone experiences with the Hirschsprung diagnostic kit from Bio-Optica (Italian company)? This kit contains lyophilizised reagenses, that have to be restored before use. It comprises the detection of AChE, SDH, ANE and NADPH. Any input is appreciated. regards Gudrun Lang

Re: [Histonet] processing cycle

Hi Allison, we doubled the times of the regular processing protocol beginning with longer absolute ethanol, intermedium and paraffin. Our regular protocol takes 13 hours and the fatty-protocol takes 17 hours. We start it at about 2 pm with endtime at 8 am. So breast tissue is mostly embedded at

Re: [Histonet] Eosinophil labelling

What about simple H? Gudrun -Ursprüngliche Nachricht- Von: Julio Benavides via Histonet [mailto:histonet@lists.utsouthwestern.edu] Gesendet: Montag, 14. September 2015 18:52 An: histonet@lists.utsouthwestern.edu Betreff: [Histonet] Eosinophil labelling Hi everyone, Is there anti