Re: [Histonet] Faded H tissue section

Hi, Don't decolonize. Soak the coverslip off and let it sit in xylene to make absolutely sure all the mounting media is off then simply re-stain it. If there is any mounting media left on it will interfere with the staining. Unless there's something wrong with either the section or the

[Histonet] Re IF staining questions

Hi, Direct IiF (a fluorescent conjugated primary) is certainly easier, but there are some reasons one might prefer using an indirect method (using a conjugated secondary). Indirect methods allow the use of a different wavelength to be used simply by switching the secondary. It is also

[Histonet] p17 mice brain sections (Alonso Mart?nez Canabal)

Hi, The thicker the section the more likely it will be to fall off. Frozen sections already love to fall off slides. You should cut them a lot thinner. 4 to 10 um should be the thickness, especially for frozens. If you want very thick sections as you describe, you would be better off

[Histonet] Detection Systems

Hi, I use polymers regularly, but keep a small kit of LSAB or the like on hand for oddball solutions like when someone gives me a biotinylated antibody or an antibody with next to no information about what it was raised in. I don't mind a biotinylated antibody all that much, but no species

Re: [Histonet] Microwaving Slides

Hi Samantha, Microwaves are terrible! I am really not a fan of them in general, but especially for drying slides, and even moreso for slides intended for IHC. There is no way to really monitor the temperature the slides get to. Sure you can get a fnacy one with a probe, but that probe

Re: [Histonet] Jones problems

Hi, I think your silver precipitation issue could be cured by the thiosemicarbizide step too. If the reaction is sped up by this, you won't need to have the slides in long enough to develop the precipitate. That said, since it is a silver stain, you still need the glassware to be acid

[Histonet] Jones problems

Hi, When I first did this stain, I had really light staining of the GBM. StainsFile has a note in the procedure that describes thiosemicarbizide. Being a complete nerd, I wanted to see another source as well. Here is a link to an article where it is used...

Re: [Histonet] Histonet Digest, Vol 194, Issue 15

Hi Betsy, I wouldn't do it. It's an unnecessary risk to let them dry out. Better to leave it in distilled water if you absolutely must. Ideal to not deparaffinize in the first place if you can't finish the stain. Dried out sections is just a bad plan. Amos Brooks On Mon, Jan 20, 2020, 1:00

[Histonet] X-gal staining

> > > Hi, I'd like to concur with Carl Hobbs. With formalin fixed, paraffin embedded tissue, beta Galactosidase is definitely the way to go. It is just an antibody so you would do it like any other IHC. Amos Hi > No replies so far so.my pennyworth. > Imho ...no > beta Gal enzyme is

[Histonet] plural fluid prep

> > > Hi Michelle, Pleural fluid lasts a fairly long time if you don't centrifuge it and leave it in suspension. Of course it will last even longer if properly fixed. I have returned to a stored (unfixed) pleural fluid for retrospective testing up to a week later. The longer you let it

Re: [Histonet] guidelines for using a microwave designated for food, but used to heat up histogel

> > Hi, There is NO reason to have a microwave in a histology lab. They don't really save any time and cause more problems than they are worth. You won't have to worry about any certifying agencies rulings about microwaves if they aren't there! Just put the Histogel in a rack in a

[Histonet] Tunel

Hi, > I hand stain this. It is a finicky test and benefits from a personal touch. When I have a ton of them, sometimes I do the enzyme on the stainer and then take it off for the primary and secondary then DAB on the stainer. Amos Message: 1 Date: Fri, 19 Oct 2018 19:24:01 + From:

Re: [Histonet] Histonet Digest, Vol 179, Issue 5

Hi, > I have a hunch that the DAB deposits you are seeing are not a DAB problem. You didn't mention how long your incubation time is. If I were to venture a guess, I am thinking there may be some evaporation of either the primary or secondary antibody or boil-over from the antigen retrieval.

[Histonet] Oil Red O

Hi, I have seen this too. I mitigate the problem by making it up in 50 ml increments and staining them flat. I draw from around the middle of the Falcon tube I make it up in. It tends to precipitate so you could filter it. Don't bother re-using the reagent. It's cheap & easy to make up. I

[Histonet] Science Haiku

Anyone that heard Science Friday recently may have heard of this. I thought it would be fun to see what others come up with. So many people think I'm an archaeologist when I tell them I'm a Histotech. This makes for a fun elevator pitch for our profession. Scientists are challenged to summarize

[Histonet] Tunel with Apoptag

Hi, I am contentedly using the Millipore (Chemicon) Apoptag IHC kit for Tunel stains. It works great... *BUT* I have been getting requests for a fluorescent tag rather than a chromogenic. The contents of the fluorescent kits are identical except for the anti-Dig. In the chromogenic kit is HRP

[Histonet] buffered zn-formalin recipe needed

Hi Johanna, The Histonet Archives are your friend. In 2009, Gayle Callis posted a great formulation that being the neo-Luddite that I am I printed and have a copy of here for just such an occasion... http://lists.utsouthwestern.edu/mailman/htdig/histonet/2009-October/046985.html Zinc

Re: [Histonet] Modified Pentachrome

Hi, I totally forgot to get the info about the safran Friday. I'll find it when I get back on Tuesday. I just didn't want you to think I had forgotten about you. A note about the safran; It is an anhydrous solution and upon making it up one would think it was goofed up because most of

[Histonet] Modified Pentachrome

Hi, I use Lugol's Iodine all the time for this. It works just fine. I do purchase certain chemicals from manufacturers like Lugol's Iodine which I get from EMS. Most of the chemicals I make up myself from powder though. I have shared the procedure with you via Google Docs. I hope it helps.

[Histonet] Crystals in PAS

Hi, If you are having trouble with the instrument and the manufacturer isn't able to fix it, just hand stain them. PAS is really not a complicated stain. Please don't just accept underperforming equipment when we are all capable of so much more. Amos

[Histonet] Mucin on PR

Hi, I'm asking this for a colleague that is experiencing this problem on formalin fixed paraffin embedded human clinical samples. She is using PR636 from Dako on a Leica Bond platform. Has anyone seen mucin or any cytoplasmic staining with Progesterone receptor antibody? Thanks folks, Amos

[Histonet] PPE/ Hair Regulations

If it is actually getting caught in things or falling in waterbaths or embedding centers and such, then it needs to be tied back. The same is true of food service. You can't risk contamination. If it is under control though, leave it alone. Don't try to use safety to enforce your personal bias of

[Histonet] Tissue Arrays

Hi, My experience with microarrays is that they are sometimes a bit complicated to work with depending on the way they are constructed and the platform they are used on. Often these slides are dipped in paraffin to help preserve them. When this happens they need considerably longer

Re: [Histonet] IHC with H & E staining

Hi, I usually try to avoid eosin as a counterstain for a DAB labeled slide because the red/pink of the eosin can obscure the rusty brown of DAB. If you really want to use it though I would suggest a *really* light eosin, perhaps even just a few milliliters in the 95% ETOH as you are

[Histonet] GATA3

Hi Lisa, Skin seemed to work fine for me. Amos On Tue, Dec 22, 2015 at 1:00 PM, wrote: > Message: 2 > Date: Mon, 21 Dec 2015 13:12:14 -0500 > From: "White, Lisa M." > To: > Subject: [Histonet]

[Histonet] VEGF

Hi, VEGF is driving me nuts. I was hoping someone might have some suggestions. I have been using clone VG1 for a while. I have never really been particularly happy with it. The labeling is not nearly as specific as I would like and there is almost always some background. If I dilute it more

[Histonet] Purple elastic

Hi, There is a reason that every manual with an elastic procedure says that the stain should be made up fresh. My guess is that the company is trying to prolong the shelf life by removing or reducing either the iodine or ferric chloride. Doing this will prevent the hematoxylin from turning

[Histonet] H2O2 blocking

Hi Gayle, Thank you again for the insight. You are a wealth of knowledge. I am also not particularly surprised that I was incorrect in my assumption about Peroxabolish. I really like to know what is in the products I am using. That's why I prefer to make up my own reagents whenever possible,

[Histonet] More on H202 issues

Hi, Peroxidase can really be a pain. If you look in the archives though (or ask her really nice) Gayle Callis submitted a recipe for a glucose oxidase for peroxidase quenching that does not include hydrogen peroxide. If you aren't really a fan of making these things up I would bet dimes to