Tylka Nematology Lab

Disinfestant solution: 0.5% chlorhexidine diacetate (hibitane)

Solution Preparation:

• dissolve 0.5 g into 100 ml sterile distilled water
• dissolve 2.5 g into 500 ml sterile distilled water

Procedure:

1. Concentrate eggs in 50-ml centrifuge tube by centrifuging at 2,000 RPM for 3 minutes.
2. Under the laminar flow hood, pour off the water (taking care not to disturb the pellet of eggs), and add the hibitane solution.
3. Cover the tube opening with a square of parafilm, shake to resuspend the egg pellet, and incubate at room temperature for 15 minutes.
4. Centrifuge the egg suspension (as in Step 1) to concentrate the eggs and pour off the hibitane solution while working in the hood.
5. Add sterile distilled water to the tube, cover the tube, and shake to resuspend the eggs, then incubate for 10 minutes.
6. Centrifuge the egg suspension (as in Step 1), pour off the water, add fresh sterile distilled water and incubate one final time for 5 minutes.
7. Centrifuge to concentrate the eggs, pour off the water and decant the surface disinfested eggs into a suitable container.