Protocol for macroscopic Sudan IV staining of arteries for fatty streaks

Prepare Solutions:

A. 1 liter of Sudan IV staining solution A:

Combine in a 1 liter Ehrlenmeyer flask:

500.0 ml Acetone

500.0 ml 70% EtOH (350mls 100% EtOH, 150 mls deionized H2O)

weigh 5 grams of Sudan IV, dissolve into acetone & EtOH, place on a stir plate for 30 minutes.

B. Decolorizing solution B:

500.0 ml 80% EtOH (400 mls 100% EtOH, 100 mls deionized H2O)

PROCEDURE:

1. Trim extraneous tissues from the tunica adventitia of the vessel

2. Drain formalin from formalin-fixed tissue and pin to paraffin tray (pinning is not necessary for small arterial rings or carotids that may be stained free floating in their original sample container).

3. Cover tissue with staining solution A for 15 minutes, rocking periodically.

4. Decant staining solution to a separate container for reuse (up to 3X) or disposal in EHS solvents container.

5. Differentiate (decolorize) the stained tissue by covering for 5 minutes with Solution B (80% EtOH) rocking periodically.

6. Decant solution B and discard in EHS used alcohol container.

7. Place under running tap water for 1 hour (un-pinned arterial rings can be placed in tissue cassettes and rinsed in a tray).

James R. Turk, 2/26/2002