Establishing an Active HIV Infection in a Rat Model

프로토콜

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. EcoHIV-EGFP [Ecotropic HIV Enhanced Green Fluorescent Protein] virus stereotaxic surgeries

1. Anesthetize rats using 3% sevoflurane. When the rats are not responsive to noxious stimuli and their reflexes are absent, proceed to step 1.2.
2. Shave the hairs from the brain region and sterilize the skin twice with 70% ethanol and a chlorhexidine-based scrub. Secure the rat in a prone position in the stereotaxic apparatus.
3. Make an incision (5-6 cm) through the skin along the scalp midline.
4. Mark two drilling positions at 0.8 mm lateral, 1.2 mm rostral to bregma. Drill a hole (diameter 0.4 mm) in each skull position.
5. Fill titrated EcoHIV lentivirus solution (1.04 × 10⁶ TU/mL) in a 10 µL injection syringe. Secure the syringe to the stereotaxic apparatus.
6. Move down the needle close to the surface of the drilling hole. Measure and move 2.5 mm in depth.
7. Infuse 1 µL of virus solution at a rate of 0.2 µL/min. Keep the needle inside of the injection area for 5 min. Slowly move the needle up until it is outside of the rat skull.
8. Suture the skin with a 4-0 silk thread.
9. Sterilize the incision with 70% ethanol once. Subcutaneously inject butorphanol (Dorolex, 0.1 mg/kg body weight).
10. Transfer the rat to a recovery chamber with a heating pad until it wakes up.

자료

Name	Company	Catalog Number	Comments
Antibiotic-Antimycotic solution	Cellgro	30004CI	100X
Cover glass	VWR	637-137
Drill
Ethicon Vicryl Plus Antibacterial, 4-0 Polyglactin 910 Suture, 27in. FS-2	Med Vet International	VCP422H
Hamilton syringe	Hamilton	1701
Iris Forceps	World Precision Instruments	15914
Iris Scissors	World Precision Instruments	500216
Stereotaxic apparatus	Kopf Instruments	Model 900
Vannas Scissors	World Precision Instruments	500086