Murine Model of Islet Transplantation: A Surgical Method to Infuse Pancreatic Islets in Portal Vein and Prevention of Postoperative Bleeding

1. Islet transplantation

1. Induce and maintain the mouse under general anesthesia with 2% isoflurane. Check for the lack of pedal reflexes to ensure proper anesthetization of the animal.
2. Shave and remove the fur in the abdomen area of the mouse.
3. Administer a single pre-operative dose of Buprenorphine (0.1 mg/kg i.p.).
4. Disinfect the surgical area with three alternating wipes of 2% iodine and 75% alcohol.
5. Perform a laparotomy with micro scissors to generate a 1-1.5 cm incision.
6. Open the peritoneal cavity with a retractor. Follow with either method A or method B as detailed below.

 Method A: (stop bleeding with gel foam, Figure 1A)

1. Mouse preparation
   1. Place a sterile gauze around the incision.
   2. Gently pull out the intestine using a forceps and keep it on the gauze.
   3. Identify the portal vein by its location and expose it well.
   4. Cover the intestine with a warm saline-wet gauze during the entire surgery.
2. Insert the islet preloaded insulin syringe needle through the portal vein near the duodenum (Figure 1B). To do so, hold the needle with the hole (bevel) facing down and position the opening surface's angle parallel to the portal vein wall before penetrating through the wall.
   1. Pull the plunger to draw some blood (20-50 μL) into the syringe to mix the islets first.
   2. Infuse the islets into the portal vein slowly while repeatedly pulling and pushing the plunge.
   3. Place a piece of gel foam (about 0.5 cm x 0.5 cm in size) to cover the injection site.
   4. Press the gel foam down with a cotton tip while pulling out the needle from the portal vein.
   5. Continue pressing on the gel for about 2 min to confirm there is no active bleeding.
   6. Rollover the cotton tip over and away from the gel foam to make sure the gel foam covers the portal vein well.

Method B: (stop bleeding with fat pad, Figure 1C)

1. Expose the portal vein thoroughly.
   1. Use two cotton tips to hold the exposed portal vein from both the left and the right sides.
   2. Identify the fat tissue pad between the duodenum and the portal vein.
   3. Penetrate through the fat pad before inserting the needle into the portal vein (Figure 1D).
   4. Infuse the islets, following the similar procedure described above in Method A.
   5. Pull out the needle while pressing down on the fat with a cotton tip.
   6. Continue pressing on the fat pad for 1 min after removing the needle.
2. After confirming that there is no bleeding from the portal vein, gently return the intestine to the peritoneal cavity in its original position.
3. Leave 0.5 mL of warm saline (36-37 °C) in the abdominal cavity before closure.
   NOTE: Warm saline facilitates post-surgery intestine movement and recovery and prevents intestine necrosis.
4. Close the muscle layer with an 5-0 suture.
5. Close the skin layer with an 4-0 suture.
6. Place the mouse in a clean cage on a heating pad until fully recovered from anesthesia.
7. Continue to provide an analgesic (e.g., buprenorphine 0.1 mg/kg i.p.) every 12 h and supplemental heat for 48 h post-surgery.
   ​NOTE: The islet transplantation procedure takes approximately 15-20 min to complete.