CRC Organoid Culture: A Method to Obtain 3D Organoids from Colorectal Cancer Cells

- Colorectal cancer, or CRC cells, travel through the circulatory system and metastasize into other organs, such as the liver, lungs, bones, and brain. To study cancer progression, we use an organoid, a 3D in-vitro model that can mimic the organotypic nature of colorectal cancer. Begin by extracting CRC tumors from an anesthetized mouse.

Mince the tumor into small pieces. Now, add the tissue pieces to a tube containing an organoid culture medium including collagenase. Collagenase breaks the tumor clumps, resulting in single-cell suspension of tumor cells.

Next, seed the cell suspension onto an extracellular medium, or ECM-coated plate, and incubate overnight. Add more cell suspension with ECM-containing medium in the wells. Upon embedment into an ECM layer, cancer cells interact with the components of the matrix. These interactions help the CRC cells grow with high efficiencies and self-organize to form organoids.

Finally, collect the organoids using a cell scraper and store them in a microfuge tube for further analysis. In the following protocol, we prepare 3D organoids from colorectal cancer to be used as an in-vitro model to study cancer progression and metastasis.

- To generate