Organoids Construction and Characterization

I. Organoid construction process:

Second, the key factors for successful organoid construction:

·  Accurate selection of tumor lesions to ensure the amount of tumor cells, freshness and cleanliness of the sample

· Accurate selection of digestion reagents and precise control of digestion time to prevent excessive digestion.

·  Collect as many single cells and small cell clusters as possible after digestion and remove incompletely digested tissue.

· Appropriate cell implantation density and high percentage of viable cells.

·  Reasonable choice of matrix gel concentration and curing method to reduce cell wall sticking after gel planting.

· Appropriate organoid culture reagents to improve the success rate of organoid construction.

· Daily monitoring of organoid growth status and timely treatment to prevent overgrowth of organoids.

III. Organoid identification:

Histomorphology

Source Tissue/Organoid

Colon cancer tumor-like organs and source tumor tissues have high morphological similarity (HE staining)

Immunofluorescence

CEA/CKA/E-cad

Gastric cancer-like organs express the same biomarkers CEA, E-cad, and CK7 as the source tumor tissue