Establishing Pure Cancer Organoid Cultures: Identification, Selection and Verification of Cancer Phenotypes and Genotypes.

Precision medicine requires in vitro models which will both faithfully recapitulate the features of an individual's disease and enable drug testing on a wide variety of samples covering the greatest range of phenotypes possible for a particular disease. Organoid technology has immense potential to fulfill this demand, but it will be necessary to develop robust protocols that enable the generation of organoids in a dependable manner from nearly every patient. Here we provide a user's guide, including detailed step-by-step protocols, to the establishment, isolation and verification of gastric cancer organoids. Selection strategies include omission of growth factors, addition of drugs, isolation of distinct phenotypes and generation of monoclonal lines. For confirmation of cancer identity, we use sequencing, drug selection, karyotyping and histology. While we specify these protocols for human gastric cancer organoids here, the methods described are applicable to organoids derived from other tissues as well.