Hello
I am trying to stain tangles in human brain frozen sections. Thioflavin T has been widely used by the people in paraffin sections, however in frozen section I am seeing lots of non specific binding. The image lights up like Christmas. Any suggestion would be appreciated.
I tried 0.5% and 0.1 % Thioflavin in 0.1N HCl.
Dear Mitesh,
guessing you are doing Fluorescence Microscopy? (or only BF-LM?)
Perhaps you have seen also a recipe for Thioflavin S staining for Senile plaques (amyloid) to demonstrate neurofibrillary tangles and senile plaques in
Alzheimer's diseased brain tissue sections at http://library.med.utah.edu/WebPath/HISTHTML/MANUALS/MODTHIOF.PDF
Also there is an article, "Binding mode of Thioflavin T in insulin amyloid fibrils" by Groenning Minna, Norrman Mathias, Flink James M., van de Weert Marco, Bukrinsky Jens T., Schluckebier Gerd, and Frokjaer Sven, in: Journal of Structural Biology 159 (2007) 483–497, stating that there are reactions for two binding site populations (but perhaps 3 binding site populations exist).
You might have also a look into:
by Ritu KHURANA et al, in Journal of Structural Biology 151 (2005) 229–238. ( and see also subchapter .
It might be that staining in frozen sections might be a bit complicated and I do apologize not to have any clue for your "lighting up like a christmas tree". Is there any specific reason you use Thioflavin T rather than Thioflavin S for localizing tangles?
Best regards, Wolfgang
Thank you for such detailed answer wolfgang. One of my colleague has worked with Th-T with paraffin sections, So I started with it. I am going to give Th-S a shot now and see if its any better. and yes you are right, the staining of frozen sections is complicated. I will look in to the articles you mentioned. Once again Thank you
Dear Mitesh, you're welcome.
Just to offer: if you can't access the above cited articles, let me know....
I'll provide later on 2 recipes out of Bancroft /GAMBLE,6th Ed(2008/2011 (originally applied to fixed / paraffin sections, Thioflavine T (but only, if you reply that you don't have these recipes = VASSAR & CULLING 1959 and BURNS et al, 1967= ph1.4 Th-T)
and - additionally - you should have perhaps a view into:
GUNTERN R, et al,1992_Experientia 48,p.8-10: (which has been described for application to cryostat sections).
This article one can find @ http://link.springer.com/article/10.1007%2FBF01923594
(only PPV on first sight, but if you click: (you find the icon above the and you are able to see the first two pages which include
i) the method as well as ii images....).
I've found other possibilities for staining tangles, perhaps more specific than Th-S (which has been said to be +/- unspecific) like X-34 (X-34, a lipophilic, highly fluorescent derivative of Congo red), cf.:
STYREN et al,
doi: 10.1177/002215540004800906 J Histochem Cytochem September 2000 vol. 48 no. 9 1223-1232, to be found as pdf free of charge/free access @:
jhc.sagepub.com/content/48/9/1223.full
Best wishes, good luck and regards, Wolfgang
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