SIRIUS RED STAINING FOR COLLAGEN

 

Materials

1.     Tissue fixation: in neutral buffered formaldehyde solution, 8-16 hours

2.     Paraffin sections

3.     Picro-sirius red solution

Sirius red F3B (C>I> 35780, Polysciences # 09400 0.5g

Saturated aqueous solution of picric acid 500ml

Add a little solid picric acid to ensure saturation ( you will see some crystal in bottom)

(Sigma #197378)

 

4.     0.2% phosphomolybdic acid

5.     Acidified water

Add 1 ml acetic acid (glacial) to 200 ml of tap water before use

 

 

 

Staining Procedure

1.     De wax paraffin sections in xylene 2x5 min and hydrate in alcohol 100% 1x5 min, 90% 1x5min, 70% 1x5min and then tap water 2x5 min

2.     Treat with 0.2% phosphomolybdic acid for 5 min

3.     Stain with Picro-sirius red for 90 minutes

4.     Rinse briefly with 0.01% HCL ( acidified water) x 2 times

5.     Dehydrate in alcohol 70% x 5min , 90%x5min, 100%x5min

6.     Clean in xylene 2x5min

7.     Dry slide and mount in Permount Medium