1. Aseptically dispense:
• one (1) 20 μL drop of H and
• three (3) 20 μL drops of ES (ES1, ES2 and ES3)
NOTE: ES1 and ES2 should be in close proximity to H (but not touching).
2. Place MII oocyte(s) (2 maximum), into H and expose undisturbed for 1 minute.
3. Merge ES1 with H. Allow spontaneous mixing for 2 minutes.
Use tip of transfer pipette to move ES1 towards H until drops merge.
4. Then merge ES2 with H+ES1. Allow spontaneous mixing for 2 minutes.
5. Transfer oocyte(s) from merged drop to ES3, and expose undisturbed for 6-10 minutes.
6. During above 6-10 minute exposure, aseptically dispense one (1) 50 μL drop of VS.
7. Transfer oocyte(s) from ES3 to VS for 30 seconds before loading.
8. Gently pipette oocyte(s) once within VS drop to ensure complete rinse in VS.
NOTE: To minimize floating, after 10 seconds pipette the specimen(s) to the bottom of the VS drop.
9. Load, seal and plunge HSV Device (per device instructions) within 80 seconds, not to exceed 110 seconds after exposure to VS.
10. Refer to HSV Device Loading Protocol diagram and product insert for detailed loading instructions.