Esophageal 3D Raft Culture Model: An In Vitro Culture Technique to Generate Human Esophageal Mucosa Model

A human esophageal mucosa model comprises human fibroblasts and epithelial cells grown on an animal-derived esophageal scaffold.

To prepare a 3D raft model of a human esophagus, begin by taking a sterile esophageal scaffold pre-treated to remove its native cells. Place the scaffold with its submucosal side up in a culture well.

Next, set a stainless-steel ring onto the scaffold. Within the ring, seed a human-fibroblast cell suspension. Subsequently, dispense a fibroblast medium around the ring and incubate.The ring confines the fibroblasts onto the submucosal surface, facilitating their adhesion.

Now, remove the ring, add fresh media to immerse the scaffold, and incubate. Fibroblasts proliferate to form a multilayer cell sheet that secretes essential cofactors.

Next, invert the scaffold, mucosal side up, and place the stainless-steel ring over it. Seed a human-epithelial cell suspension within the ring and add the culture medium. Incubate to allow the confined epithelial cells to adhere to the mucosal surface.

Then, remove the ring and add fresh media. Incubate to diffuse the fibroblast-produced cofactors through the scaffold and promote epithelial cell proliferation.

Now, using a metal grid, raise the scaffold to expose the proliferating epithelial cells to the air-liquid interface. This allows for the complete maturation of the culture into a human-esophageal model.