Indeed you can, we established a protocol about 3 years ago and have been using it with neuronal tracings. It has always worked very well in our hands. It should work with any IHC, really.

I refer you to our protocol in Article The Centrifugal Visual System of a Palaeognathous Bird, the ...

Here are some specifics regarding the buffer and how to prepare the TSA aliquots:

- Stock: 0.05M Borate-Buffer (BB) at pH8.5 (250 ml):

4.77g sodium tetraborate decahydrate (MW 381.37) + ~50ml H2Od, add 2 drops of concentrated HCl, fill to 250ml with H2Od and adjust pH to 8.5 (using HCL/NaOH).

- TSA-stock (10%): 5mg biotin-tyramide / 50µl DMSO; store as 1 µl aliquots at -20°C (can be stored for long times).

We use biotin-tyramide from IRIS Biotech, Cat# LS3500. It is quite cheap and avoids having to synthetize biotin-tyramide in the lab.

For use:

- make diluted TSA stock (0.1%): 1 µl TSA-stock aliquot + 99 µl BB

- make TSA working solution (0.0001%): 24 µl diluted TSA stock + 24 ml BB

- make 1% H2O2: 5 µl 30% H2O2 + 145 µl BB

immediately prior to use add H2O2 to 0.003%: e.g. 72 µl 1% H2O2 + 24 ml TSA working solution

- Incubate avidin-peroxidase-bound tissue in TSA working solution containing 0.003% H2O2 for 20-30 minutes (you may need to establish the precise time for your application).

It is of high importance to only use very clean (dish-washer-washed or in the best case new) well plates for doing the histochemistry. We found that re-used hand-washed well-plates can negatively influence the TSA reaction such that it does not work at all.

- Afterwards, we wash 4 times in phosphate buffered saline (30-45 minutes in total). No over night step has been necessary in our experience. Also, we have not used any stop reagents.

- Then, we continue with a second avidin-peroxidase (ABC kit) step, followed by DAB. Alternatively, we use Streptavidin-Alexa conjugates instead of the second ABC kit step, if we want to have fluorescence for double labelings with different IHCs.

If our protocol works for you, we would appreciate it if you cited our above-mentioned paper.

Please go ahead and contact me if you have any questions.

Brilliant, thanks Quirin!

You are welcome Alan! I just edited my answer to add that we use biotin-tyramide from IRIS biotech instead of synthetizing it by ourselves as in the link you posted.

I hope our protocol will be of help to you.