[Histonet] Control slides and/or blocks
I am curious to know if there is any interest in positive/negative control tissue blocks or slides (FFPE) with any number of pathogens. We are an infectious disease research center and we are always looking to expand our contract research services. We work mostly with mouse models but from time to time we also work with hamster and ferret tissues. Pathogens include SARS-COV2, Influenza virus, Zika virus, Dengue virus, as well as mycobacterium tuberculosis and many other bacterial pathogens. Thanks in advance for any thoughts on the subject! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] McNeal's Trichrome recipe
Hi everyone, I am trying to stain some tissues embedded in Osteo-bed. Does anyone have a recipe for McNeal's trichrome? Also I am having a tough time getting the sections to stay adhered to positive charged slides. Any recommendations? The sections are thin. Couple microns maybe. Thanks for any help! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] hard Tick histology
Hello, I was wondering if someone might have some expertise on how to prepare a hard-tick for sectioning in paraffin. I have been decalcifying to soften the exoskeleton but not sure if that even helps. I have used acid decalcifying but would EDTA be more appropriate? I have punctured the ticks and used vacuum for all steps (fixation, decalcification, dehydration etc). Still crunchy on the outside and soft in the middle! Any help would be greatly appreciated!! Thanks! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Large bone samples
Hi Everyone, I am looking into doing some histology (HandE) on NHP joints. Samples will be finger joints and wrist. If you have any experience with these larger bones (and soft tissue) I would be interested in any advice on general time and type of decal solution. I only need to do H&E no immuno so I think acid decal is OK. Thanks for any help!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Fluorescent assay for Eosinophils
Hi Everyone, I am looking for a specific marker for eosinophils. Came across some testing laboratories that offer Immunofluorescent assay for eMBP1 and EDN. Does anybody know of a good antibody source to label these components? Is this assay antibody based or dye based? Thanks! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Radiation sterilization and IHC
It was recently suggested to me that we might try to use our irradiator to sterilize (not fix) tissues rather than Formalin solution. Obviously we would need to validate this but has anyone tried this prior to IHC or IF? Thanks! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Sucrose cryoprotection
I'm having problems getting good cryosections of mouse lung tissue that has been formaldehyde fixed, cryoprotected with sucrose, and embedded in OCT. The lungs were fixed with 3% formaldehyde for 24 hours followed by 10%, 20%, and 30% sucrose until the tissue sank in solution. When sectioning the tissues collapse. I have tried cutting colder and cutting thicker but still getting poor sections. I embedded whole lobes of the lung individually. Any thoughts would be greatly appreciated !! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Human placental tissue
Wondering if anyone has any tips for processing human placental tissue before I try to reinvent the wheel! (formalin fixed tissue to paraffin) Thanks for any help! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Tissue clearing with scaleA2 reagent
Anyone using ScaleA2 reagent to clear tissues for imaging? I am wondering why the method requires Sucrose gradient followed by OCT embedding and freezing just to turn around and thaw and clear? ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Processor problem
I am having a problem with our tissue processor (Tissue Tek 3000). During processing the first reagent (NBF) gets pumped into the retort without any problems and the same for the 2nd reagent (70% ETOH). The Third reagent (also 70%) however, gets about half way through pumping into retort and stops. After that it reverses and pumps out reagent three, then pumps in half of reagent 2. Pumps out the half of reagent 2 and pumps in half of reagent 1. This is followed by a low fluid warning. I have removed and reinserted the reagent reservoirs and suction tube. The tubing is not caught in the handle and seems to be fine. I have created a short (5min) automatic run with 5 minutes per station (no samples) and this problem repeats itself. Has anybody had this problem? Thanks for any help! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Activated carbon
I am looking for a low cost source for activated carbon. So far I am looking at $50 per pound for 6-14 mesh. This is for a VIP 300. Anybody use a different mesh size? Thanks! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Yahoo link
Anybody wonder who has the most harzardous job to your health? We're Number One!!! http://finance.yahoo.com/news/the-15-jobs-that-are-most-damaging-to-your-health-155706120.html Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] anti-Human CD20
Thanks for all the great replies!! While I have you all worried about my cell pellets ;-) Does anyone have a good anti-HuCD20 that works well for FFPE tissues (cells). Thanks! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Embedding cells in Paraffin
Has anyone tried to embed cells grown in tissue culture? I am trying to put some tissue culture cells through same stress as tissue would go through. Fixation, dehydration, and heat. Any ideas? I could re-suspend in OCT and then fix for extended time with NBF but that doesn't quite seem fair. Thanks for any ideas! ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Frozen Embedding Medium
I recently had some tissues sent to me for frozen sectioning and staining. I was able to section them just fine and also stained them reasonably well the next day. The stains that I used were ones that I have used many times on similar sections. The stains did not seem as "crisp" as usual but the were OK. I went back to the unstained slides two days later (stored then at -20) and I had lost the ability to stain with the same antibodies. I cut some fresh sections and the staining ability was restored. I am used to keeping tissues at -20 for months and still able to stain with these markers. The tissues were embedded in a different freezing compound than I am used to using. Has anyone had this problem? I use Tissue-Tek and these tissues were embedded in Frozen section compound (w/blue dye). The formulation seems to be a bit different judging from the MSDS. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Paraplast vs. Paraplast plus
Does anyone have a preference over Paraplast and Paraplast plus for mouse tissues? Thanks!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Tissue Tek VIP 3000
In an internet search I came across a used TissueTek VIP 3000 that is in "excellent condition". This looks to be an older model and I am wondering if anyone has any experience with this instrument? Are there any issues with parts replacement? comes with one year warranty and they are asking around $8000.00. Thanks! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Human bcell and Tcell Ab
Hi Everybody! Can you guys weigh in on the best antibodies for Human B cell and Tcell staining in FFPE sections? We would like to have a fluorescent endpoint if that changes things. Thanks in advance! Hope everyone has a safe Holiday. Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Equipment service and repair
I am wondering if there are any service company's that will repair and service Leica equipment in northern New York (Saranac Lake)? I have a Leica VT 1000s that needs some attention and would need some occasional PM visits. Thanks for any suggestions! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Antigen retrieval
Does anyone have a favorite antigen retrieval method for FFPE mouse tissues that they would be willing to share? I have been using citrate buffer Ph6.0 with poor to moderate results. Thanks for any help! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] NFAT antibody
Anybody using anti-nFAT antibodies in mouse tissues or tcells? If so, could you recomend a particular antibody? Thanks! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Tissue processor
I am looking to purchase a Processor and embedding center. I have received a quote for VIP6 and a TEC5 but they seem to be a bit out of our price range. Does anyone have another Processor/embedding center that they found to be reliable? Thanks Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Baboon Tissues
Hi All, We received some frozen Baboon spleen tissue from the NIH that turned out to be very difficult to cut. I am experienced with Mouse tissues but rarely have to cut tissue the size of my thumb. The best information I could get was that the tissue was not put through a sucrose gradient but put directly into OCT. The OCT seemed to come off the blade just fine but the tissue seemed spongy and looked more like lung tissue (with many holes) once it was warmed on a slide. My first thought was that it had been put through a sucrose gradient that was not complete. I tried colder and warmer temps with thicker and thinner sections. My question list Is there anybody who has had a similar problem and do you think it is a tissue processing problem (No hope). Do you think it is a cutting problem? (only hope is for a better technician than me, maybe) I have an opportunity to get more tissues collected from a collaborator. What would be the best way to collect these tissues be? Thanks again for all your help!!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Slide stainer and coverslipper
I have been asked to provide cost estimates for a slide stainer and a slide coverslipper. I have not been given time to get a quote but thought that maybe someone would have a reasonable idea of the cost. a range of cost would be fine also. Thanks for any help! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Tissue-Tek VIP 2000-3000
I may have the chance to get a tissue processor (VIP 2000 or 3000). Are these units still serviceable? Is it hard to find parts? Are they Reliable? Thanks for any help!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Granzyme B and Lamp1 (CD107A)
Hey! Has anyone looked at Granzyme B and/or Lamp1(CD107A) in mouse cells by immunofluorescence? I know this is done for flow cytometry but thought someone may have a good method for cytospun cells. Thanks for any ideas!!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Yellow counterstain
I have seen Tissue sections counterstained with a yellow counterstain. I would like to use an Iron hematoxylin with this counter stain. Can anyone tell me what this is? Are there any tips/tricks to this counterstain? Thanks in advance! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Vibrating microtome sections
I would like cut fresh spleen and lymph node sections on a vibrating microtome. I have tried embedding in low melt agarose but the tissue seems to be too flexable to get good repeating sections (uniform thickness). When the knife does catch the cells seem to pour out into the surrounding medium. Does anybody have a suggestion or a reliable method they would be willing to share? Maybe there is a embedding medium that I am not aware of? Thanks for any Help!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] IHCWORLD orders
I have recently tried to order a product from ihcworld and had no luck at all. Has anyone had this problem or is there anyone out there that is familiar with this company/website. I think that selling products may be new to this website. Any suggestions? Thanks! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Liver tissue
I am having trouble cutting FFPE liver sections. I have embedded the tissue manually going from fomalin to water to increasing ethanol to xylene and to paraffin. I can get good sections by soaking the block with ice cold water before taking a few sections but then have to re-soak in order to take more sections. If I do not soak the tissue on the chuck the liver crumbles and falls out of the paraffin. The soaking and sectioning is very time consuming. Does anyone have an idea what might be the problem? Thanks for any help!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] course recommendation
I am looking for a conference/course to attend in 2010. I am experienced in IF and IHC and would like to learn more about any new techniques and products that might be available. Does anyone have a recommendation for a course or a conference in the US? I am not a certified HT but I pretend to be one:) Thanks for any suggestions!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Viral RNA probes
Does anyone know of a company(s) that sells/makes ISH probes to viral RNA (PR8 and or Sendai) or a company that will make custom probes. Thanks for any help!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Antigen retrieval
I am attempting to do IF on FFPE sections but the history/treatment of my tissues (mouse tissues) is unknown. I would like to recommend to our labs a strategy for tissue collection, length of time in NBF, type of antigen retrieval, and so on. Would anyone (or everyone) be willing to share their opinions on the best methods they have for maintaining the ability to do IF or IHC. Thanks for any help! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Cytoseal XYL
Has anyone compared Cytoseal 60 with Cytoseal XYL? I clear with Xylene and use cytoseal 60 (Toluene)to coverslip. Seems like it would make sense to use Cytoseal XYL (Xylene) but they only sell in a case and I can't get a sample. I would be interested to know if anyone has a preference. Thanks!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] OCT and RNA
We have taken sections from OCT embedded tissues and processed them for RNA analysis with out any problems. You should be careful to move the tissue to a solution that will protect the RNA once it has been either sectioned or thawed. Good Luck! Trudeau Institute email service will be temporarily suspended on May 2-3 to complete the joining of electrical services between the newly constructed Stafford Wing and our existing facilities. Email sent to and from all Trudeau Institute employees will be held in queue until services are restored on Monday, May 4th ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] BSL3 clean-up and Paraffin odor
We have a BSL3 laboratory with a cryostat (TB research). The cryostat user asked me for the best method to decontaminate the cryostat. My suggestion was to clean the cryostat (while cold) with 95% ethanol followed by absolute ethanol after wards. Then to defrost the cryostat and clean with a stronger disinfectant such as amphyl/lysol. Does this sound reasonable to those of you who work with BSL3 level pathogens? Is there any thing you might add to this protocol? Second, someone asked me if smelling the paraffin (not xylene) was hazardous to your health. I have never been concerned about the smell of paraffin alone but since it was asked of me I thought I would see if any of you have had to worry about this? Thanks!! Mike ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Liver
I am am having a difficult time getting immuno fluorescent staining in mouse liver. I am using markers that work side by side in lung tissue (F4/80, GR1). Does anyone have any suggestions for Immuno Fluorescent staining in mouse tissue or similar problems? Thanks!! ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
