Llewellyn's Sirius Red
for Amyloid

Solutions

Alkaline sirius red
Sirius red F3B	0.5	g
Distilled water	45	mL
Ethanol, absolute	50	mL
Dissolve the dye into the water, add ethanol and mix well. Add 1 mL of 1% sodium hydroxide.
With strong backlighting and swirling, add drops of 20% sodium chloride until a fine precipitate is seen.
Usually about 2 mL is adequate. Adding more than 4 mL causes excessive precipitation.
The solution is reasonably stable for several months, but does slowly deteriorate
Extending the staining time can compensate.

Tissue sample
5µ paraffin sections of neutral buffered formalin fixed tissue are suitable. Other fixatives are likely to be satisfactory.

Method

1. Bring sections to water via xylene and ethanol.
2. Stain nuclei with a progressive alum hematoxylin for a few minutes.
3. Rinse with tap water.
4. Rinse with ethanol.
5. Place into alkaline sirius red for 1 - 2 hours.
6. Rinse well with tap water.
7. Dehydrate with absolute ethanol.
8. Clear with xylene and mount with a resinous medium.

Expected results

• Amyloid  –  red
• Eosinophil and Paneth cell granules  –  red
• Nuclei  –  blue
• Background  –  colourless

Notes

1. Amyloid displays deep green birefringence when viewed with crossed polarisers, one above and one below the section.
2. Eosinophils and Paneth cell granules are also demonstrated. If used for this purpose the sodium chloride may be ommitted.
3. This method uses sirius red F3B. The dye sirius red 4B is not suitable.

 

Reference
Llewellyn, B.D., (1970)
An improved sirius red method for amyloid.
Journal of Medical Laboratory Technology, v 23, 308