Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.
Initiating culture from frozen vials: For a brief overview of the thawing procedure see our quickstart guide Thawing Cryopreserved Human Organoids.
Seeding density: 0.25 - 1 x 106/ viable cells in 100 µL of ECM per well of a 6-well plate. Alternatively, split at 1:2-1:4 every 7-10 days. For example, collect organoids from 100 µL of extracellular matrix (ECM) from a single well of a 6-well plate and re-seed into 2-4 wells of a 6-well plate in 100 µL ECM per well.
Media renewal: Perform a complete medium change every 2-3 days. Include 10 µM ROCK Inhibitor Y-27632 (ATCC ACS-3030) in medium for the first 2-3 days following subculture.
For a brief overview of the subculture and expansion of organoids see our quickstart guide Subculture and Expansion of Human Organoids Protocol.
For more details on the handling and culture of organoids see our methods paper in Current Protocols in Cell Biology
We recommend cryopreserving this model in ATCC Stem Cell Freezing Media (ATCC ACS-3020). For a brief overview of the cryopreservation procedure for organoids see our quickstart guide Organoid Cryopreservation Protocol.
For more details on the handling and culture of organoids see our methods paper in Current Protocols in Cell Biology.
