Figure 1: Stem cell colony morphology prior to organoid formation. Colonies should be 50%-70% confluent at the time of dissociation to produce a large number of consistent organoids. (A) Sparse, recently-seeded cultures have not yet reached log-growth phase, and will not produce many organoids. (B) Once the cells reach the proper confluence, they are ready for dissociation. It is also important that these colonies are inspected to ensure that they are clear of differentiation. (C) Colonies that are taken too far will reach over-confluence and are not recommended for organoid formation. Cultures in this state are more likely to contain differentiating cells. (D) Colony edges should be distinct, with consistent cell morphology of rounded cells in the center, and slightly elongated cells toward the edges. (E) Minimal differentiation should be present in the cultures. If larger, flattened cells are observed in the centers or edges of colonies (white arrowheads) and make up more than roughly 1% of the culture, it is recommended that pick-to-remove or additional passages are conducted to form a uniform stem cell population. Please click here to view a larger version of this figure.
