Organotypic 3D Skin Culture: A Cassette-based In Vitro Culture Technique to Regenerate Human Epidermis on Acellular Dermis

- Organotypic skin cultures help generate reconstructed skin tissue with typical epidermal differentiation and stratification characteristics. They mimic the functional and physiological in vivo tissue architecture.

To generate an organotypic skin, first, assemble a pre-cut, dead, de-epidermized dermis onto a customized cassette with its top layer facing upwards. This dermis forms the substrate to aid skin regeneration.

Invert the cassette assembly to expose the bottom layer of the dermis. Pipet an artificial extracellular matrix over the bottom layer. Allow the matrix to solidify and provide structural support to the dermis.

Invert the assembly back to its initial position. Add the desired volume of keratinocyte growth media to immerse just the bottom layer of the dermis. Add keratinocyte suspension onto the upper layer of the dermis to mimic the typical skin tissue architecture and incubate.

During culture, the growth media diffusing through the dermis supplements keratinocyte proliferation. Additionally, the disposition of these proliferating keratinocytes at the air-liquid interface promotes their differentiation. The process regenerates a fully stratified epidermis over the de