hi can i ask? first, thankyou so much u making this video. it helpss. much. when we incubate the working silver methanamine solution. can I drip that solution in to section and incubate in 65 degree celcius on hybridizer/ hot plate for 30-60 minutes?
Hi, this sounds like an interesting idea and is worth trying, however would suggest covering the slides with a lid to better maintain an even temperature and to reduce drying of solution at the edges. You could also try preheating the coplin with methenamine silver in a water bath. There will be more than one way to achieve a satisfactory result but microwave oven has always worked well for me.
@@damienharkin ah thankyou for the insight. i will try to preheat the reagent first and do drip the solution to slide inside the hybridizer with lid , its about reagent consumption matter for my customer. 😁🙏
The reaction requires heat. Theoretically you could pre-heat the silver first, then insert the slides, but I've never tried this approach. I think either way will work. The advantage of preheating is that you can mix the solution before adding the slides, thus possibly achieving a more even staining outcome. Good luck and let me know how it turns out.
Hi Damien! Thanks for your greate video! I try do do the Jones silver stain on our mouse kidney tissue. My problem is that my basement membran staining is quite weak, although Im using semicarbazid to boost the signal. Im not using a coplin jar for the silver impregnation, I put the Silvermethenamin-Borax directly on the slide an put it in the Incubator or the microwave, to save some staining solution. Do you think thats a problem?
Hi Jonas, I'm not familiar with semicarbazid, but I assume that you are oxidising your slides first using periodic acid? Apart from that, I would definitely suggest using a Coplin jar filled with solution and stain 5-6 slides at a time as shown in the video. Good luck!
@@damienharkin Thanks for your quick answer! Yes Im reducing with periodic acid. I tried 0,5% for 10min up to 1% for 30min. I dried it in the Coplin jar in a water bath. But I still have the problem, that Im not getting a good contrast. If I let incubate the Silvermethenamin-Borax for short time, the basement membranes staining is not very strong and if I incubate it for a longer time, the cells in the glom an the tubulus cells are stained as well. I fixed the tissue with 10% buffered formalin.
@@jonaseinloft2625 Over fixation might be a factor. If unable to prepare fresh specimens, then would suggest checking temperature within Coplin jar. In my experience, the basement membranes of PCT and DCT generally develop silver deposits more quickly than the glomerular filtration apparatus. If staining 5-6 slides at a time, I would normally heat in microwave for around 1 minute, then mix the solution well and wait until a dark brown ppt starts to develop across the surface of the slide and section (1-2 minutes).
thank you so much
You're welcome!
hi can i ask? first, thankyou so much u making this video. it helpss. much.
when we incubate the working silver methanamine solution. can I drip that solution in to section and incubate in 65 degree celcius on hybridizer/ hot plate for 30-60 minutes?
Hi, this sounds like an interesting idea and is worth trying, however would suggest covering the slides with a lid to better maintain an even temperature and to reduce drying of solution at the edges. You could also try preheating the coplin with methenamine silver in a water bath. There will be more than one way to achieve a satisfactory result but microwave oven has always worked well for me.
@@damienharkin ah thankyou for the insight. i will try to preheat the reagent first and do drip the solution to slide inside the hybridizer with lid , its about reagent consumption matter for my customer. 😁🙏
In the brochure of all the GMS stain kits it's said to put slides in pre-heated working solution.
Is it really important to put slides in pre-heated working silver solution?
The reaction requires heat. Theoretically you could pre-heat the silver first, then insert the slides, but I've never tried this approach. I think either way will work. The advantage of preheating is that you can mix the solution before adding the slides, thus possibly achieving a more even staining outcome. Good luck and let me know how it turns out.
Great video thanks sir
Hi Damien! Thanks for your greate video! I try do do the Jones silver stain on our mouse kidney tissue. My problem is that my basement membran staining is quite weak, although Im using semicarbazid to boost the signal. Im not using a coplin jar for the silver impregnation, I put the Silvermethenamin-Borax directly on the slide an put it in the Incubator or the microwave, to save some staining solution. Do you think thats a problem?
Hi Jonas, I'm not familiar with semicarbazid, but I assume that you are oxidising your slides first using periodic acid? Apart from that, I would definitely suggest using a Coplin jar filled with solution and stain 5-6 slides at a time as shown in the video. Good luck!
@@damienharkin Thanks for your quick answer! Yes Im reducing with periodic acid. I tried 0,5% for 10min up to 1% for 30min. I dried it in the Coplin jar in a water bath. But I still have the problem, that Im not getting a good contrast. If I let incubate the Silvermethenamin-Borax for short time, the basement membranes staining is not very strong and if I incubate it for a longer time, the cells in the glom an the tubulus cells are stained as well. I fixed the tissue with 10% buffered formalin.
But we fixed it for about 2 weeks. could this be the problem?
@@jonaseinloft2625 Over fixation might be a factor. If unable to prepare fresh specimens, then would suggest checking temperature within Coplin jar. In my experience, the basement membranes of PCT and DCT generally develop silver deposits more quickly than the glomerular filtration apparatus. If staining 5-6 slides at a time, I would normally heat in microwave for around 1 minute, then mix the solution well and wait until a dark brown ppt starts to develop across the surface of the slide and section (1-2 minutes).
Thank youuu
after-- adjust with gold , color fade away and patchy . Do you know why??
You may not have stained for long enough initially with silver. Or perhaps you have treated for too long in gold chloride.
@@damienharkin thank you.
sample is precious and can I stain that slide again?
What structures are you trying to demonstrate?
@@damienharkin renal glomerulus
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