Sample type required: Deparaffinized and rehydrated tissue section (3–5 microns) on positively (+) charged slides
Preferred fixative: 10% neutral buffered formalin (NBF)
Control: Aorta, artery, or skin
| Reagent | Time | Technical Notes |
| Verhoeff's elastic staining solution | 1 hour | Make fresh; save solution until staining is complete. Ferric chloride is required as a mordant in this (hematoxylin-based) solution. |
| Running water wash | Until the solution drains clear |
|
| 2% ferric chloride solution | Differentiate until black fibers are crisp and well-defined and the background is gray | Review slides microscopically to ensure differentiation is complete. Due to varying elastic fiber content, the control and patient slides should be checked individually and microscopically to determine accurate differentiation. If differentiation is incomplete, return to the solution for 30 seconds and repeat. If over-differentiation occurs, return slide(s) to Verhoeff solution. |
| Running water wash | Until the solution drains clear |
|
| 5% sodium thiosulfate | 1 minute | Removes iodine |
| Running water wash | 5 minutes |
|
| Van Gieson's solution | 5 minutes | Counterstain |
Post staining procedure:
The tissue section should be rinsed well in distilled water and dehydrated through graded alcohols (beginning with 95%). Two changes of xylene should follow, followed by a coverslip.
Expected results:
- Elastic fibers and nuclei - Black
- Collagen - Red
- Other tissue elements - Yellow
