Re: [Histonet] Active Caspase-3 IHC-P staining
Have a look at results in Histonet images? Good luck Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Zen light
Hi Histonet Maybe about to migrate from Axiovision to Zenlight and Zen Pro Would be grateful for any comments Thanks Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Stability Guarantee vs Expiration date
Good info I have always made 1/10 stocks of my abs ( in TBS/BSA/azide) and frozen aliquoted neat stocks. For eg: I still use a 10 yr old "Dako" anti GFAP at 1/5000 ( stock 4C 1/10) Sure, I am in research Sure, I always add a positive control Today I used a 5 yr old 1/10 4C of anti PCNA ( Abcam ab29)...worked as well as when I 1st made the 1/10 stock Sureone has to keep checking by adding controls ( a known positive like gut so that if crypt nuclei are exclusively +ve...OK) Mind you.PCNA is NOT a great prolif marker : far better to use anti Ki67 ( half-life difference) However, I wanted to see if Ki67/PCNA was good for worm Pwax sections ( former -ve, latter TOO positive) Caveat: stored positive control sections can "go off" Some call it "oxidation" of epitopes It happens to some epitopes/ags Best wishes, Histonet Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] IHC staining of cartilage
Yes...a big problem for me, tooif you are using HIER?? Cartilage and bone are major tissue retention problems I cut Pwax sections of mouse long bones/knee joints for anti GFP IHC I still get a % fall off but, using manual HIER at 90C I manage to get enough sections to get good results However, I imagine that your autoIHC stainer prob uses 90C HIER? I find Trajan 3 series slides more reliable than Superfrost Plus slides. Sure, I need to dry my slides for 24 hrs in an operating fume hood ( we can't turn ours off/down) Then I place slides in a 60C oven for 2hrs. Then dewax etc Be interesting to read of other suggestions Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Tiny folds in colon bx
Imhoyour waterbath is not hot enough The sections are not expanding Sure, if you are floating sections directly onto water bathbaaad Place your ribbon on a slide that has 20% alcohol on it Float that onto your waterbath The alcohol teases the sections nicely sono creases. NB: Leave in the waterbath until you see no creases Works for me every time NB: I do many species of gut and have no problems if.I do the above Good luck Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Detection Systems (Mac Donald, Jennifer)
As I stated: Polymer systems only work GREAT when you also use the DAB in the kit You pay dearly for it Check out my comparison in Histonet Images Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Detection Systems
Hi I'm a loner Biomed Scientist ( Histology specialist) in Research I use stABCpx-DAB regularly Yes, polymer kits are quicker and very sensitive. However, I have tested 3 popular such kits and they ONLY work well( in my hands) if you also use their DAB: I tested this by substituting my DAB for theirs ( and vice versa) and got the same signal strength as my "LSAB" std method I also used my LSAB/their DAB and got greater sensitivity So, imhoit's the DAB that gives greater sensitivity thus more dilute most expensive primary abs. Sure, if time/simplicity is of the essence...Polymer kits are the current "best" Howeverfor an extra max 60 mins...I just add Imidazole to my DAB visualisation solution after stABCpx detection That gives me identical sensitivity to polmer kits at much less cost. Sure, I can "play"...if you are in Diagnostic Histopath..you can't. Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Paraffin embedding following storage in 70% alcohol
The Most venerable Histologist ( JK) is correct...as always! No way to go from 70% alcohol directly into Pwax Have to remove ALL water before infiltrating with Pwax Yesalso need to replace alcohol with Xylene/Histoclear before infil. with Pwax. Sure, you can use Isopropyl alc to obviate xylene /Histoclear/equiv. but..Jury's out on that Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Retirement in sight!
Well, Dr Morken I do not know you but..I have read/absorbed/acted upon occasionally... your most informative Posts, over the years Your Professional History reads such a wide spectrum of interests/skills/locations! I assume that you will still be contributing to Histonet so that your valuable insights/experiences may still be available to us that have not yet retired. I trust that your Family equally enjoyed your trajectories across the World! I wish you the very best in your new "Post" Sincerely Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Protocol for DAPI staining on paraffin sections
Dear Alida my emails to your msu.montan.edu get returned to me as undeliverable I have incubated 2 dewaxed sections ( different slides) in HOECHST +/- HIER. Both stain nuclei very well. The HIER-treated section gave a brighter nuclear positivity ( so needed to lower exposure time on taking images) NSD though I can post on Histonet images, if you wish carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Re Protocol for DAPI staining on paraffin sections
Dear Alida, That is a good point. I probably have only done HOECHST on Pwax sections after they have been heat-Antigen retrieved ( HIER) However, H ( or DAPI) works on fixed cell monolayers: they don't get HIER treatment) Nor do frozen sections ( well, some do) and the HOECHST/DAPI works fine on both However, perhaps you can just take a Pwax section to water and apply HOECHST for 10 mins, wash off, mount in anti fade aq. mountant and have a look down a Fluorescence scope? If you can wait, I will check this next Monday ( put H on a non HIER'd Pwax section and get back to you by Monday later) Unless, whoever that is doing the fluorescence DAPI/HOECHST nuclear staining is actually doing it on Pwax sections that have been HIER-ed? Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6810 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re Protocol for DAPI staining on paraffin sections
Use DAPI/HOECHST at the same concentration you use for cells/FS I use Hoechst H33258 Sigma 10mg/ml soln I dilute by adding 0.5 microL to 100 microL buffer then add 1/100 to Alexa secondaries ( final 1/20K diln factor) Some incubate in HOECHST or DAPI after secondary ab incubation, for IF Sure, if only "staining" nuclei, incubate in 1/20K Hoechst in same buffer This will vary for each lab Or, buy mounting medium that contains DAPI Good luck Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Pre-made chrome alum slides source?
Hi I know that you are well aware that you can make your own, easily. (I agree re the hydrophobicity induced) I am interested to know why you do not wish to, Tim I have to admit I don't make silanised slides anymore...I buy Superfrost Plus, for example. Shame on me home-silanised are equally good, imho. Interested in differing opinions Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Evaporation in frozen aliquots
Hi Kelly If the tubes that hold your frozen ab aliquots do not have a complete sealthe water in them will lyophilise, over time. I have your problem occasionally, for that reason ( I have a couple of thousand frozen primary abs) Use snap-lid tubes ( I use 200microL PCR tubes) Hope that explains. Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] question on processing worms (1...@comcast.net)
Hi I have done several yrs of work on Eisenia fetida worms with a PhD student. I decided to do a std Pwax processing schedule. The student dissected the appropriate area then placed in 10% Formalin in PBS pH7.4 for 24hrs on a rocker ( gentle agitation) I then processed to Pwax using a std IMS/Xylene/wax on a Leica carousel ( 1hr in each station) Worked fine. Could do tinctorial and IHC no problem. Sure, many primary Abs didn't work... We also did TUNEL on them Pwax sectionsworked a treat. I may have TUNEL images in the Histonet Image archive. Good luck! NB: Sure, maybe freezing is a better way forwards. Depends. All I know is that the PhD student passed with no problems and went on to a so far succesful career in Science Industry Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Delays in tissue embedding after mouse tissue
Hi Jamie I do this often, for he same reason Ms and rat soft organs ( hollow and solid) leg bones, whole ankles, ears, cube of tissue containing cochlea, chick ribs. Once the delay was 6 months. I allowed the cassettes with tissue in to solidify ( as do you) and then leave them in a labelled box to await embedding. Sometimes the researcher cannot embed at scheduled time and I would rather fix then process than have fixed tissues sitting in 70% alcohol until they can be processed and embedded on schedule ( sure, it doesn't adversely affect, in my xp, to leave in 70% alc at 4C provided the tissues have been optimally fixed ( meaning in 10% Formalin) Obviously I strongly discourage any delay, telling them that it is deleterious Gotta keep them Researchers in order ;-) Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] human shoulder joint fixation
Jeeza human shoulder joint? Be grateful for background. Many years ago I had to fix/process to Polyester resin many monkey condylar joints ( jaw) As iterated here, I stripped all superfluous tissue off and processed the joints to resin. ( as I do now when doing Histology on ms knee joints as model for arthritisbut, using Pwax after decal) Monkey mandibles: fix in Formalin...on a rocker for 2 wks ( 20:1 Formalin:PBS) Changed twice I then took them specimens to a dedicated lab who sectioned this resin using a diamond circular saw. I was too young to follow the process further...also, in them days , Technicians did what they were told and were never appreciated. Maybe a different fix fluid will give a better morphologyJK will advise, surely! However, the project I was involved with won a prize...somewhere ( too young to be bothered) C Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Apple Green Birefringence in Amyliod slides
I have images on Histonet image archive Congo red brightfield and also fluorescence using the TRITC channel. No green birefringence images tho I find fluorescence more sensitive for Congo red However, my std is to use an anti Amyloid beta antibody. If anyone cares to look and comment.I'm always listening/reading to improve my practise/practice ( which is the korrekt word?chuckle) Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Warm formalin
You a re right, Lynette De rigueur for Diagnostic labs! My apologies for forgetting that ( I am now in research labs where...it is less restricted, unfortunately). Essential to be well-documented/adherent to SOPs. Respectful-illy Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 From: Lynette Pavelich Sent: 22 July 2020 19:19 To: Hobbs, Carl Cc: histonet@lists.utsouthwestern.edu Subject: Re: [Histonet] Warm formalin I would suggest to always refer to your reagent’s IFU insert. This will advise at what temperature you should use/store. All inspectors (CAP, JC, CLIA, etc.) will make you adhere to these specifications. Unless you do a well documented validation study that goes outside of these restrictions from the IFU that proves no patient harm, you honestly must go by the IFU recommendations. This would apply to all of our stains/reagents/solutions/antibodies. Our world is becoming more restrictive…… hope this helps, Lynette Pavelich, HT(ASCP), QIHC > On Jul 22, 2020, at 1:55 PM, Hobbs, Carl via Histonet > wrote: > > Depends on what you are doing with the sections. > IHC or just dye -staining? > Sure...too hot ( cooking) is not recommended, as stated > Also stated is that high -temp fixation may also be used with no deleterious > effects as long as the fixation time is not extended. > However, RT -ish even for a week won't be a problem...imho > Needs must? > > > > Carl Hobbs FIBMS > Histology and Imaging Manager > Wolfson CARD > Guys Campus, London Bridge > Kings College London > London > SE1 1UL > > > 020 7848 6813 > ___ > Histonet mailing list > Histonet@lists.utsouthwestern.edu > https://eur03.safelinks.protection.outlook.com/?url=http%3A%2F%2Flists.utsouthwestern.edu%2Fmailman%2Flistinfo%2Fhistonetdata=01%7C01%7Ccarl.hobbs%40kcl.ac.uk%7C2915a105cfa5436de65e08d82e6bd68b%7C8370cf1416f34c16b83c724071654356%7C0sdata=YZGiBCOwM5NmjBuxHh%2B%2FuhltOh%2F0HxJs5Kb8FaurKQM%3Dreserved=0 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Warm formalin
Depends on what you are doing with the sections. IHC or just dye -staining? Sure...too hot ( cooking) is not recommended, as stated Also stated is that high -temp fixation may also be used with no deleterious effects as long as the fixation time is not extended. However, RT -ish even for a week won't be a problem...imho Needs must? Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] An occasional blog post
Strange/coincidence that your link has my name in it, chuckle I can't access the Chicago Tribune link Or, is it a scam?? Curious-illy Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Not forgetting...
...Ham's Histology! Forerunner and...still inciteful! Set the std for rigorous Histology books. What a wonderful read, still. He talks to me Sure, Junquiera too! A moreinstant need ? Also the graphic Freeman and Bracegirdle...my students still want to take images of their images...simple( Black N white) yet...perfect! Of course they cannot. I love the simplicity of it.. Also...I cannot remember the author ...but..an orange-coloured 1st edition of Basics of chemical fixation ( not that title..I can't remember..grrr ) I love reading it tho...I don't understand most of it, chuckle ( I have it, Dear Readers...it is at work but, ...I am furloughed) Who was it written by?? Frustrated-illy Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Histology text book "bibles"
Good to read of others' xp I remember the Armed Forces book...good in parts, unlike the Curate's egg! Disbrey and Rack...wonderfully refreshing. Exceedingly well-informed person was Brenda! I reckon she swung the panel in my favour for a Histology Manager job.YEARS ago! Imho the forerunner of the best: Kiernan Sure, neck and neck with Bancroft and Stevensboth game-changers Pearce...wonderfully instructive yetdense? What about the 70's "bible"...I forget the English author..dash it! His surname begins with a C? Carleton! The book was THE text at that time ( in UK)...however, I found it rather didacticno depth I could be wrong, of course! I did learn a lot from it...then found it lacing in depth...thirst for Histo teck knowledge, chuckle It gave me a seminal Taster for more, however. Met him at many London Histology discussion meetings...at one of which he dismisssed me for using DPX mountant ( cheaper for my lab/quickersetting...close RI , I recall?) Harrumph, he saiduse natural resin: Canada balsam! Much closer RIhmm What does Prof. Kiernan think re DPX v Canada balsam?? Luverly smell..tho ( C.balsam) Does anyone use CBalsam still? Also sure, Harry Cook has a special place in my Histology career...such a humble yet...knowledgeable person. He opened up my mind to things Histological...his depth and breadth of things Histological was...humbling and mesmeric. His Carbohydrate booklet was so good for me, when I needed it! Curious-illy Always Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Fixed frozen non-paraffin mouse brain
Chuckle Thank you, John Kiernan Yes...I appreciate your filling in them gaps. I do understand/know themhaving read, over the years many of the Giants of Histology who gave me such inciteful/usable knowledge, including yourself. I just didn't want to bloat my "pennyworth" otherwise it would become several Poundsworth , chuckle. I am not worthy I recall having pp in unbuffered Formalin...we chucked it Well, in those days one emptied it down the sink! Sure, re Formic acid However, imho...that is a more theoretical problem, as Formalin was used up within a couple of weeks...in any of my Labs I used to check the pH once a week. In practice, there is much leeway in the use of Formalinprovided one is knowlegable regarding all the parameters? Sure, it is not the "best" fixing fluid butit all depends, as you intimated, on one's needs ( eg: Ultrastructural integrity v immunoreactivity) Thank you very much for your elucidations I always learn moream happy to I have the ORANGE book ( at work..I am on furlough) ...grrr...I fail to recall the most excellent author...on fixation. 1st edition I thoroughly enjoy/ed reading it It is another of my "Bibles"it expands my mind just like any Bible should...imho. A bible is like a learning curve: one must go back to it when in doubt but.go forward when in no doubt but, supported/stayed by that original knowledge Hence, I can see further by standing on the shoulders of the giants that came before me? Paraphrasingly...surely. Your opinion re the Orange-covered book ? Sure, there are many resources/publications regarding fixation...most are idiosyncraticimho. Respectfully Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Fixed frozen non-paraffin mouse brain
Prof. Kiernan, as usual, provides us all with such a depth/breadth of particular information/advice. His Histological and Histochemical methods BIBLE is still my favourite read. Respect Most researchers fix in depolymerised Paraformaldehyde because someone must've originally thought: " Hang on, if we fix in commercially bought 40% Formalin, it's got 10% methanol added ( to slow rate of formaldehyde repolymerisation) ...that will compete with Formaldehyde fixation. So, we get coagulative and additive fixation. That is not good, folkslet's get pure and use depolymerised Paraformaldehyde: pure methylene glycol polymer" I am sure Professor Kiernan can correct my inaccuracies! Anyway..I've never noticed any difference: I've worked in diagnostic labs ( unfixed frozen muscle/renal/rectal bx) and also research labs ( unfixed/ fixed frozen tissues) using both fixing solutions I have not noticed any IHC/IF difference in reactivity. Many primary abs do NOT work even with fixed/unfixed frozensome of them WILL need HIER ( at 90C rather than M/W or pressure cooker AR but, only fixed frozen of course), imho. Part of the problem is whether the antigen is linear or 3D...sorry for simplicity. I can successfully snap-freeze fixed/unfixed rat/ms brain hemispheres without using sucrose ( success measured by lack of holes at the LM level). This is because I was trained in a diagnostic lab to freeze fast but, effectively. It is a technique that requires experience for consistency of successsometimes I fail! The reason most use 20/30% sucrose is to give poor a snap-freezing technique a chance to avoid ice-crystal artefact, as stated by Kiernan). Sucrose is no panacea.technique is everything. My pennyworth-illy Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Anyone having difficulty with tissue adhesion?
I am very interested. I do IHC on Pwax sections of mouse bone, human keloid tissue ( got a project aching to start which hopefully will result in a publication) and mouse/rat contused spinal cord. All are quite difficult to keep on the slides as I have to, mostly, use HIER to get the best IHC- positive result. The best slides I have used to date are Trajan's Series 3. ( superior to commercial Superfrost Plus, homemade Silanated or double-subbed slides) However, they do not retain bone/cartilage...sure, keloid collagen is much better retained but there is still some loss Sure, I get researchers to buy only them for the tissues stated above. I will not be returning to work until 3rd Augustmaybe later. It depends...…. Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] stored in 80% isopropanol
If you have a processer using IMS ( or the more expensive ethanol) take tissues from that 80% IPA ( iso Propyl alcohol) to a fresh 80% IMS/ethanol, whichever you use as std and continue as normal ( you have no idea if the specimens were taken from water/PFA directly into 80% IPA? Thus there will be more than 20% water in there) IMS/IPA/EtOH are all compatible for Pwax processing, where a std clearing agent is used ( eg: Xylene, Histoclear) Sure, you can as suggested use increasing concs IPA and go directly into wax, bypassing the "clearing" stage but, imho longer Pwax times are needed as IPA is not miscible in Pwax unless at the high temp used.no big deal. I have used this method in the past, with success Tho I have to admit that I still for my sins, find that IMS then Xylene gives the best final Pwax block! If others disagree, I'd be grateful to know what their safer equally as good clearing agent alternatives are and the cost/block compared with using xylene. Cost/benefit? Best wishes Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Recommended thickness of Amyloid sections
Depends on what you want to do: positivity or maximise signal? I have FS and Pwax images up in Histonet image archive if you care to have a look Best wishes to all Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] too much formalin time (Curt Tague)
Hi Curt I trust you and yours are well... Imho.it will still be an excellent control, provided that you are processing it to Paraffin wax and use HIER ( antigen retrieval) before IHC/IF. Sure, as long as it has been optimally fixed ( immersion fixation with no agitation -BAD- leads to zonal variations of positivity) I have posted images on Histonet site of human tonsil that has remained in fix for a similar time before processing. Sure, I cannot guarantee all proteins will be retrievable but, in my XP, good fixation is the keynot the length of fixation. Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Glycogen detection
Q has been answered: fix in "10% Formalin" made up from concentrated liquid which has 10% Alcohol to retard reformation of paraformaldehyde polymers. Once the 40% liquid has been diluted to 4% the fixative effects of alcohol are minimised. Alcohol does not dissolve glycogen ( otherwise the dehydration stage of Pwax processing would) The Main Man JK has OK'd this. I am glad that he still posts/passes on his extensive knowledge! I can see further by having his shoulders to stand on Glycogen is still demonstrable ( with PAS) but, as JK pointed out, the Formalin will "push" the glycogen up to the plasma membrane as it fixes the proteins, giving what I learnt to call "streaming" artefact In 2014, I posted an image in Histonet Images of glycogen streaming in a FFPWS skin section stained with PAS. Have a look? Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Animal and Human Tissues
If you are referring to Pwax processing.no difference imho. I process/have processed tissues from human, axolotl, alligator, mouse, rat, marmoset, quail, chicken, pig, octopus using same schedule. Also fly larvae and earthworm You can see IHC images in the Histonet image archive Best wishes Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] How to Reduce Tissue Autofluorescence
Hi Do you refer to FIF? Or...autofluorescence? Different...as you prob know so, apologies in advance. The Wright Cell imaging Facility ( Toronto western research Inst) pdf: very informative FIF: I have tried Glycine, Ammonium chloride and Na-borohyd. I got best results with Glycine Howevernone are great. Multi spectral imaging is the best way forwards butexpensive. Sure, in Lambda mode using Zeiss Zen on a confocal gives good results ( tho I have forgotten how to do it, sigh) After setting up you hit the fluorescence you don't want...hit the one you wantif the wavelengths are different, what you don't want you eliminate, electronically. Vectorlabs sell an excellent kit for FIF elimination...sure over-expensive, given the ingredients. We found that dilution of their working reagent by x2- x4 gave best results As good as multispectral imaging, imho. Autofl of lipofuscin is supressed using Sudan BlackB. Good luck Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] X-gal staining
Hi No replies so far so.my pennyworth. Imho ...no beta Gal enzyme is inactivated by std Pwax processing. So, when you apply the X-gal substrate and chromogen solution to Pwax sections, it will not give you a positive blue final reaction product If you want to detect beta Gal in FFPWS and visualise with DAB, you will have to use an anti beta Gal antibody. Finding an anti beta Gal enzyme antibody, that works in FFPWS +/- AR is not so easy. Good luck Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 From: histonet-requ...@lists.utsouthwestern.edu Sent: 08 January 2020 18:00 To: histonet@lists.utsouthwestern.edu Subject: Histonet Digest, Vol 194, Issue 4 Send Histonet mailing list submissions to histonet@lists.utsouthwestern.edu To subscribe or unsubscribe via the World Wide Web, visit https://eur03.safelinks.protection.outlook.com/?url=http%3A%2F%2Flists.utsouthwestern.edu%2Fmailman%2Flistinfo%2Fhistonetdata=01%7C01%7Ccarl.hobbs%40kcl.ac.uk%7C5189a7f8df3b4f65b02708d79465e645%7C8370cf1416f34c16b83c724071654356%7C0sdata=c06fBHXWIVgPOnNXhSw7mEFHHvThteuUFXxa5UYSlCI%3Dreserved=0 or, via email, send a message with subject or body 'help' to histonet-requ...@lists.utsouthwestern.edu You can reach the person managing the list at histonet-ow...@lists.utsouthwestern.edu When replying, please edit your Subject line so it is more specific than "Re: Contents of Histonet digest..." Today's Topics: 1. X-gal staining (Margaryan, Naira) 2. (no subject) (Waitts, Celeste) -- Message: 1 Date: Tue, 7 Jan 2020 18:21:31 + From: "Margaryan, Naira" To: "'histonet-request (histonet@lists.utsouthwestern.edu)'" Subject: [Histonet] X-gal staining Message-ID: Content-Type: text/plain; charset="us-ascii" Hello histonetters, I have request from a scientist to perform X-gal staining on the formalin fixed paraffin-embedded kidney. I was wondering if there is any possibility to do this staining on FFPE tissue as well as if there any possibility to perform a peroxidase DAB staining for the X-gal? If so, may I ask you for a detailed protocol to perform this staining. Any input and explanation why it is impossible is appreciated. Thanks in advance, Naira -- Message: 2 Date: Wed, 8 Jan 2020 16:46:18 + From: "Waitts, Celeste" To: "'histonet@lists.utsouthwestern.edu'" Subject: [Histonet] (no subject) Message-ID: <131bf77daa5d4ba08f9076d9afbf1...@exchnode1.local.centrastate.com> Content-Type: text/plain; charset="us-ascii" Question VIP6SHORT BIOPSY RUNUSING CLEARITE ANYTHING Celeste Waitts Histology Supervisor Centrastate Medical Center Freehold, NJ 07728 (732-303-5071) -- Subject: Digest Footer ___ Histonet mailing list Histonet@lists.utsouthwestern.edu https://eur03.safelinks.protection.outlook.com/?url=http%3A%2F%2Flists.utsouthwestern.edu%2Fmailman%2Flistinfo%2Fhistonetdata=01%7C01%7Ccarl.hobbs%40kcl.ac.uk%7C5189a7f8df3b4f65b02708d79465e645%7C8370cf1416f34c16b83c724071654356%7C0sdata=c06fBHXWIVgPOnNXhSw7mEFHHvThteuUFXxa5UYSlCI%3Dreserved=0 -- End of Histonet Digest, Vol 194, Issue 4 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Delay in embedding?
I agree with replies/statements 3, 4, 5 Anyone who states differently have not experienced this contingency. Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] How to prevent bubbling in pressure cooker for immunohistochemistry procedure
Hi Subash Imho your sections are not sufficiently adherent to the slide(s) It is not M/W "bubbling" that causes your problem If you use Superfrost Plus/lab-coated Silane or even double subbed slides ( thanks Ole!) AND dry your sections onto the slides efficiently, you will not get section-loss. NB: SF+ are expensive: if you cannot afford themexperiment with in-house Silane ( APES) or subbing, to coat your IHC slides. If you use the latter methods, make sure you slides are very clean. Unless the tissue is very fibrous/bone/cartilage/severely inflameddifficult to keep sections adherent. In the latter case, Trajan 3 series can help much more than std coated slides. What tissues are causing you problems? For most soft tissues I use Superfrost Plus slides, dry the mounted sections under a fan O/N ( at least one hour) For my peace of mind I then put the slides into a 60C oven for 30-60 mins before dewaxing. Please post your opinion/results so far? Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] biopsy staining
Hi Are you processing to Pwax? If so, I stain my mouse DRGS ( very small) using Haemalum. 5 mins max ( I use Gill's I) then blue in alkaline tapwater for 10 mins before Pwax processing Stain from Formalin or alcohol but.rinse in dist water to remove before staining. The "blue" stays nicely in the tissues, for embedding : sure, I then subject the sections to HIER without pretreatment as the citric acid treatment gets rid of any dye. If doing dye-staining I will acid-alcohol treat the sections before staining. Works well for me. Good luck Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re guinea pig IHC
Ms Ruegg, as usual , gives excellent advice: avoid HRP. Use Alk phos? Block end. alk phos by using levamisole. However, Ms Shivers does not state the fixation status of her FS. Is the Gpig tissue perfused-fixed then frozen orfrozen as unfixed...then fixed? Alsowhy 0.3% H2O2? Use 3%kill the enzyme, not feed it? NOT aq for FS Make up in IMS ( 74OP) No tissue disruption However: you state that you get loadsa bubbles...so what? Is your section still attached to your slide? Can you then carry out successful IF/IHC? If yesno problem. Sure, there's the argument that using a coagulant ( alcohol) in block is a No No I never had a problemprovided that the Formalin fixation was sufficient for unfixed crosections ( 15 mins) I do STILL severely dislike FS ( sure, I spent many years in Diagnostic Histopath doing Operative FS) Why not use Pwax sections, Ms Shivers? Curious-illy Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 From: histonet-requ...@lists.utsouthwestern.edu Sent: 02 February 2019 18:00 To: histonet@lists.utsouthwestern.edu Subject: Histonet Digest, Vol 183, Issue 2 Send Histonet mailing list submissions to histonet@lists.utsouthwestern.edu To subscribe or unsubscribe via the World Wide Web, visit https://emea01.safelinks.protection.outlook.com/?url=http%3A%2F%2Flists.utsouthwestern.edu%2Fmailman%2Flistinfo%2Fhistonetdata=01%7C01%7Ccarl.hobbs%40kcl.ac.uk%7C1546c2da69de40ae400f08d689398cc4%7C8370cf1416f34c16b83c724071654356%7C0sdata=gEf%2FzAdfyRdZG0CI%2Bty4tXrOxlk5NwyD4O9qxP8DZ0E%3Dreserved=0 or, via email, send a message with subject or body 'help' to histonet-requ...@lists.utsouthwestern.edu You can reach the person managing the list at histonet-ow...@lists.utsouthwestern.edu When replying, please edit your Subject line so it is more specific than "Re: Contents of Histonet digest..." Today's Topics: 1. Re: cytology listserv (Webster, Thomas S.) 2. Re: guinea pig IHC (Patsy Ruegg) -- Message: 1 Date: Fri, 1 Feb 2019 18:28:30 + From: "Webster, Thomas S." To: "'histonet@lists.utsouthwestern.edu'" Subject: Re: [Histonet] cytology listserv Message-ID: <93fc6a1cc62f41f5ad5890786ab04...@crh.org> Content-Type: text/plain; charset="us-ascii" Haven't seen any cytology listservs except the one for members of the ASC. There are some cytology facebook pages where you could get questions answered. This Histonet listserv is very informative. CONFIDENTIALITY NOTICE: This e-mail message, including all attachments, is for the sole use of the intended recipient(s) and may contain confidential and privileged information. You may NOT use, disclose, copy or disseminate this information. If you are not the intended recipient, please contact the sender by reply e-mail immediately. Please destroy all copies of the original message and all attachments. Your cooperation is greatly appreciated. Columbus Regional Hospital 2400 East 17th Street Columbus, Indiana 47201 -- Message: 2 Date: Fri, 1 Feb 2019 18:39:54 + From: Patsy Ruegg To: Jan Shivers , histonet Subject: Re: [Histonet] guinea pig IHC Message-ID: Content-Type: text/plain; charset="iso-8859-1" Especially a very blood tissue like GP spleen. Patsy Ruegg, HT(ASCP)QIHC Ruegg IHC Consulting 40864 E Arkansas Ave Bennett, CO 80102 H 303-644-4538 C 720-281-5406 prueg...@hotmail.com From: Patsy Ruegg Sent: Thursday, January 31, 2019 11:51 AM To: Jan Shivers; histonet Subject: Re: [Histonet] guinea pig IHC In my experience it is not that GP have a higher peroxidase level, it is frozen sections in general that cannot be blocked with h202, unless they are fixed for a long time in formalin. What are others experiences with h202 blocking on frozen sections. I always used an IHC detection system that did not require h202 blocking for frozen sections. Patsy Ruegg, HT(ASCP)QIHC Ruegg IHC Consulting 40864 E Arkansas Ave Bennett, CO 80102 H 303-644-4538 C 720-281-5406 prueg...@hotmail.com From: Jan Shivers Sent: Tuesday, January 29, 2019 12:58 PM To: histonet Subject: [Histonet] guinea pig IHC Has anyone ever performed IHC on frozen sections of guinea pig tissue? I am experiencing an enormous amount of bubbling when doing the peroxidase blocking step, even though I'm only using a 0.3% concentration of H2O2. And when I say 'enormous', I mean it's like continuous champagne bubbles rising out of the tissue, even after 20 minutes in the H2O2 solution. I can't find anything in the literature that mentions guinea pigs having a higher peroxidase content in their tissues. Thanks for any help that anyone can provide. Jan Shivers Senior Scientist IHC/Histology
Re: [Histonet] Frozen sections and cold acetone
I have always "fixed" in RT acetone for 10 mins Have compared 0-60 mins/4C to RT acetone Lower temps just limit the rate of reaction, imho. I note "nuclear streaming" when I use acetone at any temp/time. Imho, acetone is not an effective fixativeit's a delipidiser. So, give it 10 mins at RT. If anyone thinks it's effective because it is a dehydrant….well, one re- hydrates the section to carry out IHC/ICC/IF. It works very well for only a few abs. When testing new abs out on FS/cell monolayers ( unfixed) I always compare Formalin, Acetone, Methanol and methanol/acetone 1:1 Imho Best wishes to a great site/membership. Carl ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Elephant Tissues
If your tissues are embedded in Pwax and do not cutsorry! You have XP with many tissues so, as corroborated, elephant should be also OK You do not state what the elephant tissue is. If it is skinmaybe a problemI have problems with human keloid skin samples. I am currently assessing Trajan 3 slides ( for adherence problems with SuperfrostPlus sides) As stated: no specie is any different. Well, yes, there are differences. For eg: worm Pwax sections are not as adherent as Ms/rat...etc. I do not know why Sigh Great forum! Carlos ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Unstained slides
Hi Depends on what you mean by cryosections. Unfixed/fixed? Stored at RT, 4C, -20C, -80C. Stored dry or in glycerol So many variables! My opinion is to store blocks and cut sections as required. Least variables. Sure, one loses some tissue everytime one cuts anewa good thing. It IS complicated so, a project has to be thought out well in advance. Stimulating post Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Unstained slides
I agree: cut only the sections needed. Saves space. Sure, you lose several sections of tissue when cutting more sections. That is acceptable because, if this "oxidation" theory is true, then the initial sections will be no good. However, careful organisation of exptl procedure before actual cutting will work very well. Actually, not many Ags get "oxidised"for eg: I can demonstrate GFAP in sections that are a year old ( sure, they are stored at 4C just in case) These slides are used for Yr 1 BSc practicals and are consistently positive. Nobody knows why some Ags ( and not others) lose their antigenicity, imho Oxidation is a vague reasoning. Just like nobody really knows why HIER works: however, I am in the dipole moment school of thought, rather than the Ca++ skool Sure, in Formalin-fixed specimens. Curious-illy Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] osmium tetroxide
Please ask why Osmium is required? Good advice given so far. If it is to "stain" for myelin, surely an anti Myelin antibody should be used? Visualise using a complementary Fluorophore to that which is indicated but not explained, in your request? Good luck Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Alcian Blue digested w Hyaluronidase - any alternatives?
Vielen dank, Michael Very kind of you to send this to me. Alcian blue is indeed an interesting dye I thought that it was no longer manufactured? I am wrong, obviously. I have used the CEC method many years ago when analysing articular cartilages and it is good to see that it is still a valid method to differentiate the various acidic mucins Alles Gute carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 From: Dr. Michael Gudo (Morphisto GmbH) Sent: 24 August 2018 11:22:55 To: Hobbs, Carl Cc: histonet Subject: Re: [Histonet] Alcian Blue digested w Hyaluronidase - any alternatives? Hello Carl, Alcianblue is q quite interesting stain! Maybe this selection of a presentation helps you to find a suitable method. I attach you a PDF-document to this email. The slides are in german, but I think you will understand them anyway. Kind regards Michael Am 23.08.2018 um 20:32 schrieb Hobbs, Carl via Histonet mailto:histonet@lists.utsouthwestern.edu>>: Perhaps Alcian blue at pH3 ( all acidic mucins) on one slide/section and Ab pH1 ( highly sulphated mucins only) on a sequential slide? Sure, instead use Alcian blue CEC method? I will be interested to read other responses to your request. Mucin-illy carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu<mailto:Histonet@lists.utsouthwestern.edu> http://lists.utsouthwestern.edu/mailman/listinfo/histonet MORPHISTO GmbH PD Dr. phil. nat. Michael Gudo Weismüllerstr. 45 60314 Frankfurt am Main Telefon: 069 / 400 3019 - 62 Telefax: 069 / 400 3019 - 64 E-Mail: michael.g...@morphisto.de<mailto:i...@morphisto.de> Internet: http://www.morphisto.de/<https://emea01.safelinks.protection.outlook.com/?url=http%3A%2F%2Fwww.morphisto.de%2F=01%7C01%7Ccarl.hobbs%40kcl.ac.uk%7C1ba1d9476b684e63735808d609ab976a%7C8370cf1416f34c16b83c724071654356%7C0=pq042s3UPpkmLVabW2yshvdlZg7q53mm9G2jWIBX%2BqY%3D=0> Vertretungsberechtigter Geschäftsführer: Dr. Michael Gudo Registergericht: Amtsgericht Frankfurt Registernummer: HRB 74954 Umsatzsteuer-Identifikationsnummer gemäß § 27 a Umsatzsteuergesetz: DE243397199 Diese Nachricht ist ausschliesslich fuer den bezeichneten Adressaten oder dessen Vertreter bestimmt. Beachten Sie bitte, dass jede Form der unautorisierten Nutzung, Veroeffentlichung, Vervielfaeltigung oder Weitergabe des Inhaltes der Email nicht gestattet ist. Sollten Sie nicht der vorgesehene Adressat dieser Email oder dessen Vertreter sein, so bitten wir Sie, sich mit dem Absender der Email in Verbindung zu setzen und anschliessend diese Email und saemtliche Anhaenge zu loeschen. This message is exclusively for the person addressed or their representative. Any form of the unauthorized use, publication, reproduction, copying or disclosure of the content of this e-mail is not permitted. If you are not the intended recipient of this message and its contents, please notify this sender immediately and delete this message and all its attachments subsequently. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Alcian Blue digested w Hyaluronidase - any alternatives?
You are right! Thank you for correcting me. Surely sialomucins will be positive for PAS and H.acid will be negative? Curious-illy Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 From: Mac Donald, Jennifer Sent: 23 August 2018 19:52:52 To: Hobbs, Carl Cc: histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Alcian Blue digested w Hyaluronidase - any alternatives? The hyaluronidase is digesting out hyaluronic acid, a component found in connective tissue mucins. This reaction is used to differentiate connective tissue mucins from epithelial mucins. Hyaluronic acid and epithelial sialomucins will give the same staining results, both positive for alcian blue pH 2.5 and negative for pH 1.0. -Original Message- From: Hobbs, Carl via Histonet Sent: Thursday, August 23, 2018 11:32 AM To: histonet Subject: Re: [Histonet] Alcian Blue digested w Hyaluronidase - any alternatives? Perhaps Alcian blue at pH3 ( all acidic mucins) on one slide/section and Ab pH1 ( highly sulphated mucins only) on a sequential slide? Sure, instead use Alcian blue CEC method? I will be interested to read other responses to your request. Mucin-illy carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu https://emea01.safelinks.protection.outlook.com/?url=http%3A%2F%2Flists.utsouthwestern.edu%2Fmailman%2Flistinfo%2Fhistonetdata=01%7C01%7Ccarl.hobbs%40kcl.ac.uk%7Cc3bfd2048e1b4508a26b08d60929a33c%7C8370cf1416f34c16b83c724071654356%7C0sdata=CTVwv5AklHrbGKIoml5W4NVoYJ7Skdfro9F%2FXE0uuWM%3Dreserved=0 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] cadaver tissue processing for histology
Rinse in dist water ( or tap water) x3 over 30 mins No big deal to do this but nice to get rid of extra smells in your processing reagents ( depending on where you start processing: I start in 90% alcoholsure, I ask for recd specimens to be rinsed in water then transferred to 70% alcohol before I will process them) Hopefully, you don't want to do IHC as inclusion of Glut. is counterproductive. Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Alcian Blue digested w Hyaluronidase - any alternatives?
Perhaps Alcian blue at pH3 ( all acidic mucins) on one slide/section and Ab pH1 ( highly sulphated mucins only) on a sequential slide? Sure, instead use Alcian blue CEC method? I will be interested to read other responses to your request. Mucin-illy carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Unstained slides - how long are they good for?
I agree with Jamie Only a few Ags are "oxidised" ( that's the term used, I recall) but, don't let it be YOUR protein of interest. If you really are concerned, cut fresh sections and immunostain along with your stored sections. Imho: cut as few sections as you need. Store any unused at 4C Best wishes Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Calretinin for Hirschsprung's
When I was doing them biopsies ( pre-operative) to establish status, we used a std histochemical method on frozen sections of biopsy material ( visualising acetylcholinesterase activity). It used Osmium to enhance /darken positivity so, NOT recommended these days. I recall somebody leaving the lid off the Osmium ( stored at 4C)…..the fridge plastic lining was BLACK the next day! So, any antibody that identifies nerve fibres and ganglion cells should be equally effective, using IHC/IF? Sure, interpretation is critical ( identifying presence/absence of ganglion cells and "significant" increase in lamina propria of nerve fibres as compensatory mechanism) There must be an internationally recognised/accepted protocol? Which technique does Great Ormond Street Hospital/St Thomas' Hospital use, these days? Curious-illy Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Bouin postfixation
Hi Pablo What is the name, source and catalogue number of your antibody? ( maybe you do not need to fix in Bouin's fixing fluiddo you know, as a fact, that Bouin's fix is better?) Imho, once you have fixed in Formalin ( depending on the time in fixing fluid), you cannot successfully "refix" in another fixative. Also: "embedding": do you mean in OCT or Pwax? Curious-illy Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 From: histonet-requ...@lists.utsouthwestern.edu Sent: 04 June 2018 18:00 To: histonet@lists.utsouthwestern.edu Subject: Histonet Digest, Vol 175, Issue 3 Send Histonet mailing list submissions to histonet@lists.utsouthwestern.edu To subscribe or unsubscribe via the World Wide Web, visit https://emea01.safelinks.protection.outlook.com/?url=http%3A%2F%2Flists.utsouthwestern.edu%2Fmailman%2Flistinfo%2Fhistonet=01%7C01%7Ccarl.hobbs%40kcl.ac.uk%7C3fc139402a6e4a115ffa08d5ca3f90d2%7C8370cf1416f34c16b83c724071654356%7C0=lkgAB3JHrCmogfTpMGuXOjWVnYxZqGLoE7DKkcRxsD0%3D=0 or, via email, send a message with subject or body 'help' to histonet-requ...@lists.utsouthwestern.edu You can reach the person managing the list at histonet-ow...@lists.utsouthwestern.edu When replying, please edit your Subject line so it is more specific than "Re: Contents of Histonet digest..." Today's Topics: 1. Bouin postfixation (SANCHEZ QUINTEIRO PABLO) -- Message: 1 Date: Sun, 3 Jun 2018 18:00:43 + From: SANCHEZ QUINTEIRO PABLO To: "histonet@lists.utsouthwestern.edu" Subject: [Histonet] Bouin postfixation Message-ID: Content-Type: text/plain; charset="iso-8859-1" Dear histonetters, I have to do immunohistochemistry with an antibody that works better in Bouin fixed samples. But I am afraid that my samples -just unprocessed- are in formalin. Do you think that it could be help to transfer them to Bouin 24 hours and then 70 ethanol and embedding? Thanks in advance Pablo Sanchez-Quinteiro Lugo. Spain -- Subject: Digest Footer ___ Histonet mailing list Histonet@lists.utsouthwestern.edu https://emea01.safelinks.protection.outlook.com/?url=http%3A%2F%2Flists.utsouthwestern.edu%2Fmailman%2Flistinfo%2Fhistonet=01%7C01%7Ccarl.hobbs%40kcl.ac.uk%7C3fc139402a6e4a115ffa08d5ca3f90d2%7C8370cf1416f34c16b83c724071654356%7C0=lkgAB3JHrCmogfTpMGuXOjWVnYxZqGLoE7DKkcRxsD0%3D=0 -- End of Histonet Digest, Vol 175, Issue 3 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Congo Red
That's a good point, Bob (mouse Amyloid controls for human sections) The amyloid produced by these various mouse models (TASTPM, TGs) is human amyloidas you know. I suspect that it wouldn't be accepted as a positive control ( I don't know) because it isn't pathologically produced It is genetically induced ( as you know) I have posted images of IHC- and Congo red-positivity ( the latter also under fluorescence microscope) of amyloid in mouse brain here ( Histonet Images archive) Respectfully Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] processing Dosophilla flies
Hi Processing them , how? To Pwax? To snapfreeze? To resin? Kind regards Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Histology Hacks
I thought Histonet was a primary source of "hacks"? Obviously...not. I have many hacks, as many others do, too... Have to admit that Histonet is rather Conservative/lazy Eg: In Imagesthere's a potential MASSIVE catalogue so, why do you not add your images to the Histonet archive??? Histonet image archive should be better/bigger than ANY other Dye/IHC/IF archive? It should be, as is HPA/Gensat/Allen , a REFERENCE Sigh, it is not. yet. ?Or , I am wrong? Please tell me Carl ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Beta-Amyloid antibody for IHC
If you use clone 6E10 it will demonstrate beta Amyloid only, after Formic acid AR After Citric acid HIER, APP will also be demonstrated Imho Check out Biosource ( Invitrogen) 44-344 and 44-348? Something about x-reactivity between Abeta 1-42 and Abeta 1-43? Or not. I'd be interested to know if this is valid Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] logging in to Histonet Images
Hi My apologies for using this wayI don't know how else. I have a new PC ( lost all my links) and forgot how to log in/haven't got the link , then upload images/edit them All I can do is upload images. I'd be most grateful for link so I can edit my posts. Thank you! carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] What happened to Ultraclone/PGP9.5?
Ultraclone THE supplier of PGP9.5 for many years ! Respect to them/him/her/LTB No longersigh. Ultraclone: "legendary"...chuckle Sureother Suppliers now offer anti PGP9.5 abs Why then do they still call it "PGP9.5"?? We now know what the protein is. PGP: "Pretty good protein" I recall? Why the 9.5? Usually a clone but, but Ultraclone ab was a rabbit poly. One could only order via fax, I recall. Isle of Wight location: Rossiters Farm. Any more info re Ultraclone's history would be most appreciated. Curiously Carl ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Disposal of Bouin's Solution
"I agree it is very dangerous. When I discovered a dried bottle of picric acid someone had donated to the school right after I first came to the university. I had to call the fire department and the bomb squad came to get it. They took it to the firing range and blew a huge sized whole in the ground, this was a small bottle of picric acid. Of course we made the local news, but get rid of it if possible or keep it under water as Fred says. " Chucklewhy didn't you just wet it? It would only be "dangerous " if you hit it with a hammer The explosive used was FAR bigger than the bang from one bottle of Picric acid Overkill. The story about a factory involved in Picric acid explosion involves several magnitudes higher than any volume stored in a Lab In the volumes used in a laboratory.not really a problem unless used TOTALLY incompetently/maliciously Sureif incompetence is thereyou will get a bang akin to that one used to be able to buy from a Joke shop. FORMALIN is toxic, poisonous...it KILLS Don't use it! Do we stop using it? Hmmm In a well- run laboratory, I am FAR more concerned regarding the toxicity of stepping outside of my lab into the street where them cars are continuously spewing out far more toxic fumes. ALL OF THE TIME Yepalso the particulate matter from them car tyres that we breath in Does anyone regulate this? Nope! So, all you car drivers that are concerned re them small amounts of laboratory Picric acidthink again. Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Disposal of Bouin's Solution
Not a disposal query. "There are substitutes for it for most purposes." Bob, I am interested in "most" and "substitutes" In what situation does "most" not apply? What substitutes? For example, I read that tissues are fixed in Picric acid-containing Formalin for IHC/IF. Why? What's the logic/rationale? Always listening/reading/asking/learning Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] CD8 for rat - also happy to take human
I am very interested to have your information! Thank you Best wishes Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] ultraclone RA95101 PGP 9.5
Hi Ultraclone no longer make this Ab, sigh Always a strange Supplier.Rossiters Farm/Ultraclone's setup always made me think of Royston Vasey However, I have replaced it with Abcam's ab108986 See images of results on ms, rat and human FFPWS on Histonet Images website I think it's excellent ( I use it mainly to see/count C fibres in mouse epithelium so, it's GOT to be good). Abcam also do an excellent Ms monoclonal ( see Histonet image site also) Sure, DAKO's ( Agilent) anti PGP 9.5 is also good for human but, I have not tested it on ms/rat. Or Cedarlane: https://www.cedarlanelabs.com/Products/Search?location=1=Description=ULTRACL=PGP+9.5=0=0 which offers alternatives. Good luck! Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Immunohelp website?
Hi I am grateful for the opportunity to load IHC/IF etc images on this website. Thank you for creating it. Shame that more Immunopeople do not upload their images. Why do they not do this? Perhaps Histonet would be interested in expanding? Thus , any chance of you Main guys creating an Immunoportal -equivilent?? A great website, devised /run by the excellent Hogne! Hogne is long gone, sadly ( sure, only re his website: I am sure he is alive and wellI wish him well) Hogne had a Q and A , a methods and an image module. Histonet jest has a general Q forum? Which is invaluable buttoo broad? To be sure...I could be missing the point Respectfully, carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] PGP 9.5 Ab x-reactivity
Hi I'd be grateful if anyone can confirm whether or not Dako's Z511601-2 or Leica's ( Novacastra's) PGP9.5-L-CE anti PGP 9.5 antibodies work on mouse/rat Formalin-fixed Paraffin wax sections. Thank you Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] DAB neutralization and disposal
Store it and give it to your Chemical people ( sensible) orfollow John Kiernan's most excellent advice that he garnered ( in Histonet archives) I wonder what people do, who use polymer DAB kits? I reckon they chuck it down the sink.do the Suppliers of such kits advice appropriate disposal? Curiously Carl ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] head kidney issues
It's endogenous biotin, Tyrone. Make your own biotin blocking kit or buy RTU kit. Also, endog. peroxidases are eliminated by 3% aq H2O2 ( saves methanol) for 10mins at RT Also, don't bother with permeabilisation.you've done all that by processing the tissue to Pwax Also, use 10% Formalin. That's what I use for my zfish Pwax IHC All suggestions: imho. Have a look in Histonet image archives for IHC Pwax sections of z.fish Best wishes Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] PFA Fixed tissue not sticking to slides?
No...never add sucrose to "PFA" when fixing tissues for freezingor any other time. Fix, then sucrose- immerse until tissue sinks to bottom of receptacle if you can't snap-freeze effectively Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] PFA Fixed tissue not sticking to slides?
Hi Jenny. Can you use Pwax embedding as an alternative? H will always be better on Pwax sections. IHC will also be in many instances. What abs are you probing with? Single/double-labelling? Yesyou are fixing for too long ( for frozen sectioningfor Pwax you can fix your specimens for a week and still not suffer IHC-problems: after HIER they will or will never work, depending upon ab) PFA( Formalin) fixed sections are well-known for not being as adherent as unfixed frozen material. Why not freeze unfixed? No sucrose required ( all that does is allow for slow freezing without, in most cases, formation of ice-crystals: good snap freezing does not need sucrose pretreatment) If you are after good morphology then Pwax is the way to go, imho. If you have to cut PFA-fixed then frozen tissues, after mounting on slides stick them in a 50C oven for 2hrs. Use Superfrost Plus slides. Then use or, store at 4C for a week or freeze. For the latter 2, ALWAYS take out of fridge/freezer and immediately place under a fan/operating fume hood. Do not allow condensation to form. Orplace them immediately into that same 50C oven for an hour. Do not do this to unfixed frozen sections! Sure, check out EC's excellent advice ( as always) re slides. Interestedly, Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Porcessing FFPE tissue without alcohol??
Thanks, Liz. If you look at fat all the time, using Osmium.you then are not sure if you use K-dichromate? I am a tad confused Alsowhy not trim the block too much? Best wishes, Carl NB: Rene stated that I wouldn't be able to use any other fat stains...that's the point, Rene. I don't need any other. I commit to Osmium. Yep...there are many variations for fat staining. Imho...most are histochemical mythology. Osmium "stains" fat...histologically. As do the conventional fat-soluble dyes when using frozen sections. I would only listen to alternatives/disagreements from JKiernan. I am still waiting for the next "generation" Histo person. Cook, Kiernan.who are the other Seminals?? Enquiringly Carl From: Elizabeth Chlipala <l...@premierlab.com> Sent: 26 March 2016 15:58 To: Joanna; Rene J Buesa Cc: Hobbs, Carl; histonet Subject: RE: [Histonet] Porcessing FFPE tissue without alcohol?? We use osmium post fixation to look at fat all of the time in mouse liver, nerve and muscle samples. It works well, sample size needs to be thin, samples are friable and can crack easily. We use a specific procedure for this it includes potassium dichromate I think, I'm at home but on Monday I can send the reference. One more thing don't trim into the block too much. Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Premier Laboratory, LLC PO Box 18592 Boulder, CO 80308 (303) 682-3949 office (303) 881-0763 cell (303) 682-9060 fax l...@premierlab.com Ship to address: Premier Laboratory, LLC 1567 Skyway Drive, Unit E Longmont, CO 80504 From: Joanna via Histonet [histonet@lists.utsouthwestern.edu] Sent: Saturday, March 26, 2016 9:20 AM To: Rene J Buesa Cc: Hobbs, Carl; histonet Subject: Re: [Histonet] Porcessing FFPE tissue without alcohol?? How about Sudan Black stain? > On Mar 26, 2016, at 4:32 AM, Rene J Buesa via Histonet > <histonet@lists.utsouthwestern.edu> wrote: > > The only problem I see is that the fat will be preserved, as you wrote, as a > black osmium oxidate but you will not be able to use any "standard" fat > stain; otherwise it will work.René > >On Friday, March 25, 2016 2:41 PM, "Hobbs, Carl via Histonet" > <histonet@lists.utsouthwestern.edu> wrote: > > > > > Fix the tissue in Formalin, wash well in dw, then place very small pieces > into Osmium tetroxide solution ( std soln for TEM post-fixation) > Processing to Pwax as usual. > Basically, you will see lipids as black ( oxidised osmium) > That's the only way to demonstrate solvent- soluble lipids, using Pwax > processing. > Sure, there are caveats but, in the main...it will be Ok, imho. > I invite comments as I may be doing exactly this very soon, to count > myelinated nerve fibres in a sciatic nerve. > > > > > > Carl Hobbs FIBMS > Histology and Imaging Manager > Wolfson CARD > Guys Campus, London Bridge > Kings College London > London > SE1 1UL > > 020 7848 6813 > ___ > 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 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] Porcessing FFPE tissue without alcohol??
Fix the tissue in Formalin, wash well in dw, then place very small pieces into Osmium tetroxide solution ( std soln for TEM post-fixation) Processing to Pwax as usual. Basically, you will see lipids as black ( oxidised osmium) That's the only way to demonstrate solvent- soluble lipids, using Pwax processing. Sure, there are caveats but, in the main...it will be Ok, imho. I invite comments as I may be doing exactly this very soon, to count myelinated nerve fibres in a sciatic nerve. Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Human nuclear antibody MAB1281
I had/have the same problem with that ab on FFPWS +/- AR Also negative, under same conditions: Serotec's 6970-1257, Abcam's ab5675 I use 2 abs on hu teratomas in mouse host: Takara clontech STEM121 antibody Abcam anti MTCO2 ab110258 Images to be seen on Histonet Images site Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] What has happened to Histonet Images site?
Curiously, Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] IgG4
Images of IgG4-positive cells in tonsil FFPWS on Histonet Images website Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] IHC on old slides
Amplification is the only way. Possibly, tyramide amplification will help. It will cost you to buy a kit...sure, make your own butyou have to weigh up pros/cons Sureyou will have to play with variables. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] F480 antibody
Doesn't work in Pwax sections...for me. Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Re. Decalcification with formic acid sodium
Mouse knee joints: done lots of decalcified FFPWS for assessment of articular cartilage degeneration models. See Histonet images for a Tol blue image. Decal in 10 % EDTA for 3 days on a rocker at RT. Sure5days if you are worried. No difference in Immuno-reactivity, imho. If you want to use buffered Formic acid, use Formic acid; sodium formate. Use of citric acid with Formic acid does not make a buffer. It's just mixing two relatively mild acids. However, I am sure that Prof Kiernan can further enlighten us. Respectfully, Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] tissue fixation-formaldehyde concentrations
I have to agree with the student, John. Sure, he is coming from ignorance ( not a bad situation: naivety is not a fault... we all are/were there at some point;-) Sure...I agree with you re the using of the word Paraformaldehyde as a fixative I often sigh when used. However, differential fixation ( zonal fixation) has always been an issue. We often see the zonal effects of this? Particularly in lymph nodes ( parenchymatous tissues) Obviously, when immersion fixing. Most often ...NOT with agitation. Respectfully, Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] RE..... murine CD4, CD8 and CD68 for FFPE tissue
I use the anti CD68 clone FA-11 ( obtained from Abcam). It works very well on frozen sections but, I have been unable to get any positivity on Pwax sections ( using +/- Citric acid pH6 HIER) Carl ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re: thanks (Truscott, Tom)
All the very best, Tom. I checked out where you workedseems an essential service that can't be replaced. Sureprivatised? I don't know you butlosing/discarding essential skills is awful. I hope you are as successful in the future as you have been in the past. Respectfully, Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 From: histonet-boun...@lists.utsouthwestern.edu histonet-boun...@lists.utsouthwestern.edu on behalf of histonet-requ...@lists.utsouthwestern.edu histonet-requ...@lists.utsouthwestern.edu Sent: 01 January 2015 18:03 To: histonet@lists.utsouthwestern.edu Subject: Histonet Digest, Vol 134, Issue 1 Send Histonet mailing list submissions to histonet@lists.utsouthwestern.edu To subscribe or unsubscribe via the World Wide Web, visit http://lists.utsouthwestern.edu/mailman/listinfo/histonet or, via email, send a message with subject or body 'help' to histonet-requ...@lists.utsouthwestern.edu You can reach the person managing the list at histonet-ow...@lists.utsouthwestern.edu When replying, please edit your Subject line so it is more specific than Re: Contents of Histonet digest... Today's Topics: 1. thanks (Truscott, Tom) -- Message: 1 Date: Wed, 31 Dec 2014 19:42:53 + From: Truscott, Tom ttrus...@vetmed.wsu.edu Subject: [Histonet] thanks To: 'histonet@lists.utsouthwestern.edu' (histonet@lists.utsouthwestern.edu) histonet@lists.utsouthwestern.edu Message-ID: 9ef5279ebdfe6e4fb6605e8f183a0027d69c6...@cvm76.vetmed.wsu.edu Content-Type: text/plain; charset=us-ascii Many thanks to all who contribute and keep the histonet running. I'm working my last day here at USDA-ARS ADRU within WSU which has been a very rewarding and enjoyable occupation. I was honored to be on 14 publications in 12 years with a few more in the works. Due to Federal budget constraints, they don't plan on replacing me with another BS An. Sci. ASCP HT. Happy New Year to you all! Thanks again Tom T= Thomas C. Truscott -- ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet End of Histonet Digest, Vol 134, Issue 1 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] FoxP3 and Geminin Immunohistochemistry
I have no xp re Geminin but, I have posted images of FoxP3 in FFPWS on the Histonet Images website. The two abs work well for me. I used a homemade std Citric acid HIER solution. Looks like you have choices, given Katy Milne's response! Good luck. carl ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] RE: lab math: ihc dilution
?Imho, the Ab concentration is irrelevant. I consider dilution factors more important. You can have a 1mg/ml Ab concyet it may not work in any particular application. Think affinity/avidity.these are critical. Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re: can you defrost and fix muscle tissue?
Your project reads ill-thought out! I agree with Elizabeth re defrosting. Should be fine. Yes, if your freezing is slow, you will most likely have ice-crystal artefact. You will see this in the Pwax sections. You have large pieces of muscle. Good that you fixed for longer. Imho, if you are not requiring Immunostaining, the longer the fixation...the better ( within reason) I would assume your problems are a result of UNDER processing. Muscle pieces that size, I would always process to Pwax over 40hr schedule. What was your processing schedule? I often Pwax process 1x1x0.5cm pig skeletal muscle and have no problems cuttingnope, no ice involved. Just a sharp knife and, a gentle rub of the block face every section, using 20% alcohol -damped soft tissue. If you are examining blood vesselsyou HAVE to perfuse-fix, imho.for best results. When you get damn fine sections: Do an HE and an HVG, initially. As Elizabeth statedstick to Formalin fixation. NB: PFA is not a fixative. It is a polymer of methylene glycol and is...a powder! When dissolved it forms multiple forms : methylene glycol is the monomer. What you buy when you get 40% Formalin is methylene glycol in solution ( Formaldehyde gas dissolves in water to ~ 40% saturation). Dilute that 1/10 and you get 10% Formalin ( same as 4% PFA) The ONLY reason them Molecular biologists/chemists decided to use PFA is cos they have NO idea re Formalin/methylene glycol fixation! Neither do I, chuckle. Well.the reason they decided to use Paraformaldehyde powder is because the immediate product is , more or less, pure methylene glycol. In bought 40% Formalin, they add a small amount of alcohol to retard formation of Formic acid and polymerised methylene glycol( PFA). So.them geezers thought: OMG...alcohol! Noit will change the chemistry of fixation! Surethem geezers NEVER bothered to look at most clinical labs in the world who buy/use ~40% Formalin to make up their 10% Formalinwith great success!! Imho, nobody has produced a convincing raison d'être for adding PBS. It is added because we think of physiologyadd PBS to a fix and we get ...better fixation. ChuckleFormalin ( methylene glycol) KILLS cells. Adding PBS...hmmm. I still do.chuckle. Curiously, as ever. carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Image uploading
I make mistakes when I upload images So, I'd be most grateful for an edit function to be included. Thanks for the opportunity to share images. Sure, the excellent Immunoportal was The Best...I hope that Histonet can equal that website quality? Carlos Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Submitting images
If possible, can an edit facility be included? Sometimes I do a typo error Mea culpa-ly. Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re: Posting Images for Histonet (Marvin Hanna)
Looks a great idea; simple to use. Thanks. I used to post images on the impressive Immunoportal website but, that is sadly no more. Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re: Alizarin Red on undecalcified bone
Strip off all soft tissue ( if you don't need it) Fix ( in Bancroft and Stevens the method uses alcohol but, we have used Formalin pH7). Follow the Tripp and MacKay method. Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] rat's hind paw skin
If they are perfused, you don't need to worry about stopping them from curling ( they will be fixed/rigid). Get as large a piece as possible: preferably two pieces. (The second piece can then be further fixed for 2hrs before placing into 30% sucrose until the specimen sinks, then snap-freezing. If you don't have a cryostat, forget this.) The piece for Pwax: place in a cassette that you know has holes too small for the specimen to fall thro. I use those plastic mesh inserts: helps to keep tissue flat and safe. They fit nicely into std plastic cassettes. If your current processing protocol is good for your specimens, it will be fine for rat footpad skin. ( if you are going to do HIERyou will have to use charged/coated slides, of course.) You will lose some integrity of the dermis ( collagen will be disrupted), unless you play around with sub-boiling temps. Thanks and.good luck. Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] rat's hind paw skin
Barry has jolted me into noticing...thanks. I occasionally prepare such specimens for Pwax for IHC to detect PGP9.5-positive nerve fibres in the epidermis. Sostd 10% Formalin fixation ( us PFA if you must : I fix for 24 hrs minimumno problem if longer fixed ) and processing. I get the researcher to place the fresh dissected footpad sample onto filter paper, so it fixes flat ( otherwise it might curl). Doesn't matter if it comes off after fixation.it will stay flat during processing. Sure, if you are concerned that it will not stay flat, use plastic mesh inserts to sandwich the skin...place them in std plastic processing cassettes. If you are looking for melanin, you will see it on a HE, if it is present. Appreciate any comments on this, Barry. Always grateful for further insights. Carl ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re: Cleaning stainless steel base molds
I put mine in my 60C oven for a day or longer. Then re-use themnever have a problem... for 20 yrs. Sure, place on absorbent paper in most appropriate orientation. Sure, when I was in a lab where I was not in charge...I had to use the recycle routine on a processor or...manual xylene clean. Not necessary. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Fixing whole testes
I would immediately bisect them longitudinally with a sharp blade, then continue to fix for at least two days ( length of fixation time is irrelevant for anatomical purposes). You should have done much earlier ( sure, you run the risk of loss of anatomical relationships but.I can't see what choice you have, given the size of the tissues. I am always interested in others' views. Yes and, fix on a rocker roller at RT. A faster penetration fixing fluid would have been betteryou would have to test a few, under your conditions. NB: Formalin penetrates a 1cm cube of gelatin 250mm in 24hrs so, penetration through a dense heterogeneous tissue, especially the initital penetration of the T albuginea, will be much slower, allowing for autolytic ( necrotic, really, as the tissue is dead) changes towards the centre of the tissue. Best wishes, Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Fixing whole testes
My sincere apologies...I left the HTML option on, for the 1st post. I would immediately bisect them longitudinally with a sharp blade, then continue to fix for at least two days ( length of fixation time is irrelevant for anatomical purposes). You should have done much earlier ( sure, you run the risk of loss of anatomical relationships but.I can't see what choice you have, given the size of the tissues. I am always interested in others' views. Yes and, fix on a rocker roller at RT. A faster penetration fixing fluid would have been betteryou would have to test a few, under your conditions. NB: Formalin penetrates a 1cm cube of gelatin 250mm in 24hrs so, penetration through a dense heterogeneous tissue, especially the initital penetration of the T albuginea, will be much slower, allowing for autolytic ( necrotic, really, as the tissue is dead) changes towards the centre of the tissue. Best wishes, Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
RE: [Histonet] Bone/cartilage/epithelial tissue stain
A HE will allow you to identify those three tissue types, in normal tissues, of most organisms ( certainly in avian embryos, imho), IF you know your Histology. There is no stain/demonstration technique that will substitute for this knowledge, imho. Sure, a special stain will help to confirm the tissue type to a high degree. I usually start with staining the section with Alcian blue pH3, then place in Hx ( I use Gill's 2sure, they state you must use Iron Hx...not necessary, for me) for 10 mins. Then place in Van Gieson's solution until what I know is collagen type I is red. I hope this adds to previous excellent posts. Best regards, Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Invitrogen A6455 rabbit anti-GFP dilution for
IF using Alexa - conjugated secondaries ( indirect IF) should give you the same results as a std stABCpx-DAB detection, for any given dilution of primary Ab. However, if you are using enhanced DAB.sure, the primary Ab will have to be less dilute when visualising with an indirect IF method. So, do one run with your primary at 1/5K, 1/10K, 1/20K ( on three sequential sections, of course) ...detecting with, for eg, Alexa anti Rabbit 594 at 1/1K. Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
RE: [Histonet] Processing thin slices of mouse cerebellum
http://www.cellpath.us.com/magento/cellpath-on-line-shop/specimen-processing-and-embedding/small-biospy-processing/cellsafe/cellsafe-biopsy-capsule-blue.html I find them very good for brain slices. There is a deeper and a shallower space, depending on which way they are folded. Carl Hobbs FIBMS Histology and Imaging 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.uk ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] CD45 Ab
I need an Ab against CD45 ( LCA) that works in mouse FFPWS. I would be most grateful for a solid recommendation ( you know it works, rather than what is said on spec sheet) Thank you Carl Carl Hobbs FIBMS Histology and Imaging Manager Wolfson CARD Guys Campus, London Bridge Kings College London London SE1 1UL 020 7848 6813 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
re: [Histonet] Pen-Fix
If you want this recipe for fixing tissues for FFPWS, then performing Immuno: doesn't work +/- any AR, for me. Nice for morphology but, no better than many other precipitant+additive fixing fluids. Imho, it's like many other fixes that don't really work in a BIG way. Well, imho, if they all did...we would all be using them? Well, we all use HIER... THAT was a Paradigm shift! NB: a fixative is a chemical: dissolved in a solvent, it is then called a Fixing fluid. Pedantically, Carl Sent from my PC ;-) ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
re: [Histonet] Immunofluorescence on FFPE skin
To me, the critical data is: which Abs do you want to use on FFPWS of skin, please? If you do chromogen IHC on your skins already, with them Abs, you can do IF. Sure, autofluorescence can be a problem but..not that big a problem. Interestedly, Carl NB: that article is..strange. Carl Hobbs FIBMS 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] Re: zebrafish and IHC
Good point by Teri Johnson. I don't decalcify, thus my results may well be compromised by this procedureand, sure, the bone is disrupted. I am only interested in CNS/PNS so, I don't worry about bone/cartilage disruption. I fix in std 10% NBF. Imho, it is better to overfix rather than to worry about optimal short fixation, if you are using HIER. Utilitarily, Carl Carl Hobbs FIBMS 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] Re: zebrafish and IHC
Should be no difference ( no tricks) Have a look here for some images: http://www.immunoportal.com/ Carl Hobbs FIBMS Histology and Imaging 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.uk ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re: zebra fish experts......
Why are you fixing ? Try unfixed frozen? Or, why not Pwax embedded? They cut well in Pwax. Sure, you may have to polish the cut surface with tissue damped in water/20% alcohol to prevent cracking/curling just before each section is cut. Or, after fixation, place in 30% sucrose until kidneys sink. Store at 4C until you freeze ( if not immediately). You will have to try several approaches until you find the correct one for your Lab. Good luck. 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] Re: tricks about antibody generated in human on human
Good advice given. Biotinylated primaries to be followed by stAvidin-488 or stABCpx-DAB, for eg. Or, if biotinylated primary gives good result with above but, at low dilution factor, use tyramide amplification? Human- on- human can be just fine, depending on what proteins you want to demonstrate and, in what tissue. What tissue are you working on? What tissue prep ( Pwax/frozen)? You using patients' serum containing auto-antibodies? Or, do you have a human that you are injecting human Ags into, then bleeding that person so you can harvest Abs? Scary! NB: You suggested diluting out your secondary...not good as you will just lower your specific signal as well, imho. Curious Carl Carl Hobbs Histology 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] Re: holly nuclei bat man
Hmmm...I have raised this issue beforeno answer. All answers to datepure conjecture! Lol...if you meant Holly.rather than Holy or even Holey...nice one, Batwoman! Caped Crusader. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] CD64 Pwax Ab
Hi, If anyone knows of an anti CD64 Ab , that is specific for FCGR1 ( not x-reacting with II, III for eg), that is known to work on Human FFPWS, I would be most grateful for Supplier and #. Thanks. Carl ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
