In vitro culture of intestinal organoids for cell death assessments

In vitro cultured organoids exhibit highly similar structures to their corresponding tissues and organs. Intestinal organoids derived from intestinal stem cells faithfully recapitulate the key characteristics of native intestinal epithelium, including morphological features, physiological functions, and personalized responses to specific stimuli. This protocol focuses on the isolation of intestinal crypts from murine intestinal tissues and the subsequent establishment of stable intestinal organoid cultures in vitro. Furthermore, propidium iodide (PI) staining coupled with high-content imaging analysis was employed to characterize organoid cell death. This optimized protocol provides a robust platform for high-throughput screening of intestinal disease-related cell death factors, thereby facilitating mechanistic investigations into intestinal pathogenesis.