Decellularized Porcine Esophageal Scaffold Preparation: A Technique to Generate Acellular Scaffold from a Pig Esophagus

To prepare the decellularized pig esophageal scaffolds, begin by using scissors and a pair of tweezers to cut open a pig esophagus longitudinally. Next, wash the esophagus in a 180-milliliter sterile pot containing 100 milliliters of sterile PBS for 10 minutes with gentle shaking to remove any debris.

While the tissue is being rinsed, spray a cork dissection board with 70% ethanol. When the cork is dry, pin the esophagus onto the board with the mucosal side up. Now, use sterile tweezers to grasp the mucosal surface at one end of the esophagus and pull the tissue away from the board to separate the mucosa from the underlying submucosa.

Then, using a sterile scalpel blade, dissect the esophagus longitudinally along the lamina propria, removing the pins as the dissection progresses to allow the mucosa to be lifted away. Discard the underlying submucosa, and use a scalpel to cut the remaining mucosa into 5-centimeter square pieces. Wash the pieces in 150 milliliters of sterile PBS with gentle agitation as just demonstrated.

After five minutes, transfer the tissue into a 180-milliliter pot containing 150 milliliters of 1 molar sodium chloride solution supplemented with antibiotics at 37 degrees Celsius. After 72 hours, transfer the tissue into 150 milliliters of sterile PBS, at which point the epithelium will have begun to visibly separate from the underlying tissue.

Use sterile forceps and a scalpel blade to remove the detaching epithelium, and place the remaining tissue in a fresh pot of 150 milliliters of sterile PBS with three five-minute washes with gentle agitation. The tissue must now be stored in glycerol for a minimum of four months to fully sterilize the scaffold.