If I wanted to stain for Hyaluronan within a tumour sample slide, what would be the best way to do this?
I’ve used this protocol. http://www.ihcworld.com/_protocols/special_stains/hyaluronidase_alcian_blue_ellis.htm
Alternatively, you can use hyaluronic acid binding protein. https://www.amsbio.com/hyaluronic-acid-binding-protein/
https://www.lerner.ccf.org/bme/hascall/documents/PEGNAC%20-%20DAB%20HABP%20Staining%20Protocol%201-14-16.pdf
Ellen A G Chernoff , thanks for that. I like the idea of using alcian blue. Has this been well validated previously or have any of your publications used this method?
I use it in the opposite way, to detect sulfated proteoglycans, but with the hyaluronidase it is a standard tumor pathology stain since the 1970’s. The papers usually cited are: https://www.nature.com/articles/218099a0 and. Article Chemical and histochemical characterization of mucopolysacch...
There are kits commercially available.
(most recently I have been doing plain acidic Alcian Blue stain for sulfated PGs: Article Meningeal Foam Cells and Ependymal Cells in Axolotl Spinal C...
)
Ellen A G Chernoff , thanks for those journal links. Usually, I would have used HABP for looking at Hyaluronan but I think using alcian blue sounds feasible. Do you personally think it would be suitable for staining tissue samples looking at HA?
Yes, I think it is acceptable. The dye pH and enzyme quality are crucial. Millipore bought the Japanese company that makes very high quality enzyme. (There’s a lot of cheap stuff used to reverse cosmetic fillers for plastic surgeons)
Hi Joseph Tonge
According to my knowledge, biotinylated HABP is a better way to visualize HA. There are more controversies when it comes to Alcian Blue staining. Unless you are doing both and comparing them to each other, I suggest sticking to bHABP.
Hi Hossein Jahedi
Thanks for that. I originally thought about using HABP - but wanted something perhaps easier (?). I am going to analyse paraffin fixed tissue sample, and also have read about "biotin-TSG-6-ΔHep-Fc" as an alternative to HABP. Any idea which is better? (assuming you've used both before)
Thanks
Good job for doing a good search of the literature Joseph Tonge
Alcian Blue is easier but if you're specifically looking at HA and not other GAGs/mucins, bHABP is the one to go with!
"biotin-TSG-6-ΔHep-Fc", aka HTI-601, seems to be similar to bHABP for detecting HA. However, I'm not sure if Halozymes is still selling it, you might want to check. There are studies that show HTI-601 is more sensitive, but I have not personally used it and hence not completely sure. I think you might want to make that decision based on availability, price, ease of protocol, etc.
Many Thanks Hossein Jahedi ,
Yes having looked, unlikely to be able to easily source HTI-601 in UK. I think I will stick to using both alcian blue and HABP. Hopefully at least one technique will let me see the HA and its varying molecular weight.
Thanks!
Joseph Tonge No worries!
Just regarding the molecular weight that you mentioned, none of he method, as stains, will not allow you to observe molecular weight.
I'd like to indicate this paper with great considerations concerning the fixation technique. Good luky.
Article Patterns of Hyaluronan Staining Are Modified by Fixation Techniques
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