Gladstone Institutes Histology and Light Microscopy Core Saf-O staining Protocol

Saf-O is a method used to detect cartilage and mucin on both paraffin or frozen tissues

Tissue: Formalin fixed paraffin embedded tissue or cryosections (with less resolution and quality), sectioned at 2-10m

Note: Bake slides prior to staining, cartilage tends to come off the slide – be gentle

Controls: cartilage and bone

Solutions: 0.02% Fast green Stable for 1 yearFast green 0.1gdiH2O 500ml

1% Acetic acidGlacial acetic acid 10mldiH2O 1000ml

1% Safranin O Stable for 1 yearSafranin O 5gdiH2O 500ml

Procedure:1. Dewax and rehydrate to water2. Stain in Weigert’s working solution 10 min3. Rinse gently in diH2O 10 min4. 0.02% fast green 1min Do not rinse5. 1% acetic acid 30sec Do not rinse6. 1% Safranin O 10 min Do not rinse7. 95% alcohol 10 dips8. 95% alcohol 1 min9. 100% alcohol 2 x 1 min10.Xylene 3 x 1 min11.Mount in DPX (Gurr)

Results: Cartilage (Proteoglycan specific - Orange to redBackground - Green

Reference:http://www.urmc.rochester.edu/musculoskeletal-research/core-services/histology/protocols.cfm

26 Dec 2012 Saf-O staining Page 1 of 2

Images:

Cartilage punch SafO 2x

Cartilage punch SafO 4x

Cartilage punch SafO 4x

26 Dec 2012 Saf-O staining Page 2 of 2