Protocol for serial organoid formation assay using primary colorectal cancer tissues to evaluate cancer stem cell activity

Summary: Organoids can enable the study of solid tumors initiated from a single cancer stem cell (CSC) ex vivo. We describe a serial tumor organoid plating protocol using primary colorectal cancer (CRC) tissues as a rapid and cost-efficient approach to evaluate the impact of therapeutic interventions on CSC functions. We detail the isolation of primary colorectal CSCs, organoid embedding, serial passaging, and CSC-related analytical techniques.For complete details on the use and execution of this protocol, please refer to Masibag et al. (2021) and Bergin et al. (2021).