Long-term culture-induced phenotypic difference and efficient cryopreservation of small intestinal organoids by treatment timing of Rho kinase inhibitor

Figure 5 Enhanced recovery of long-term cryopreserved intestinal organoids was dependent on the timing of Y-27632 treatment. Undissociated organoids (P3) were treated with 10 μmol/L Y-27632 at the indicated time points. A: Representative morphology of organoids cultured on day 7 in the presence of ENR medium after thawing organoids cryopreserved for 3 mo in the presence of 10% DMSO. Scale bars, 200 μm. B: Quantification of recovery from cryopreserved organoids was performed using MTT assays. The data are shown as means ± SDs of triplicate experiments (^bP < 0.01, two-way analysis of variance with Dunnett’s T3 tests) and normalized to untreated samples. MTT: Methyl thiazolyl tetrazolium; ENR: Epidermal growth factor/Noggin/R-spondin1.