Generation of a Three-dimensional Full Thickness Skin Equivalent and Automated Wounding

The overall goal of this procedure is to build up a three dimensional full thickness skin equivalent, which resembles natural skin and to generate precise and reproducible wounds under sterile conditions using an automated wounding device. This is accomplished by first embedding primary human dermal fibroblasts in a collagen one hydrogel. Next primary human epidermal keratinocytes are seated on the top of the fibronectin coated hydrogel.

Then the model is cultured under submerged conditions, and then under an air liquid interface, a three-dimensional full thickness skin equivalent is formed, composed of a dermal component and a multi-layered epidermis. Finally, a standardized wound is generated in a sterile environment. Ultimately, immunohistochemical staining and microscopy is used to assess the histological structure of the wounded full thickness skin equivalent.