(A) The expression of the intestinal oligopeptide transporter (PEPT1) and major ATP-binding cassette (ABC) transporters ABCB1 and ABCG2 in hESC-derived day 50 gut organoids was analyzed by qRT-PCR and compared to that in the healthy adult small intestine. Statistical analysis was performed using a t test or a Mann-Whitney rank-sum test (**P < 0.01). Data are shown as means (%) ± SEM; n = 3. (B) Gut organoids were treated with fluorophore-conjugated dipeptide β-Ala-Lys-AMCA with or without angiotensin-converting enzyme inhibitor captopril (top row and bottom left). The gut organoids clearly showed dipeptide uptake (top right) inhibited by captopril (top left). Gut organoids are visible in the the bright-field image (top left), the fluorescence image (top right), and the combined bright-field and fluorescence image (bottom left) (n = 3 for each group). Schematic diagram of the peptide absorption assay (bottom right). Scale bar: 200 μm. (C) To quantify the uptake of β-Ala-Lys-AMCA, the AMCA-related signals in gut organoids cultured with or without 10 μM, 100 μM, and 1 mM of captopril were observed using a fluorescence microscope BZ-X710 (Keyence) equipped a top-stage incubator (5% CO₂ at 37°C) (images on the right), and the fluorescence signal intensity was quantified by using Hybrid Cell Count BZ-H3C (Keyence). The results did not show dose dependency. All sample images were recorded under standard conditions. Each concentration assay was performed with 3 biologically independent replicates.