Figure 1. Primary Organoids Grown in IntestiCult™ Organoid Growth Medium (Human) are Fully Mature After 10-14 Days in Culture

Primary organoids were cultured from human colonic biopsy samples and grown in IntestiCult Organoid Growth Medium (Human). Organoids were imaged after (A) two days, (B) six days, (C) eight days and (D) ten days growth.

Figure 2. Organoids Grown in IntestiCult™ Organoid Growth Medium (Human) Display Markers of Human Intestinal Epithelial Cells

Immunofluorescence of organoids grown in IntestiCult™ Organoid Growth Medium (Human) showing colocalization of (A) DAPI, (B) EPCAM and (C) Ki67. (D) A merged image shows the position of actively proliferating (Ki67+) intestinal stem cells within the epithelial layer (EPCAM+).

Figure 3. Forskolin-Induced Swelling of Organoids Grown in IntestiCult™ Organoid Growth Medium (Human)

Organoids were treated with (A) 5 μM Forskolin or (B) DMSO and organoid area was measured at 0 minutes and 120 minutes. (C)Forskolin-treated organoids increased in size 33.5 ± 3.8% compared to 7.5 ± 0.8% for DMSO-treated organoids.

Figure 4. IntestiCult™ Organoid Growth Medium (Human) Supports the Growth of Organoids in Multiple Extracellular Matrices

Intestinal organoid cultures were prepared in IntestiCult™ Organoid Growth Medium (Human) and plated in (A) Matrigel® Growth Factor Reduced Basement Membrane Matrix (Corning® catalog # 356231), (B) Geltrex® LDEV-Free Reduced Growth Factor Basement Membrane Matrix (Gibco™ catalog # A1413202), (C) Cultrex® Reduced Growth Factor Basement Membrane Extract, Type 1 (R&D Systems™ catalog # 3433-005-R1), and (D) Cultrex® Reduced Growth Factor Basement Membrane Extract, Type 2 (R&D Systems™ catalog # 3533-005-02). Organoid cultures are imaged at the end of passage 4. All four extracellular matrices supported robust growth of human intestinal organoids. Scale bars = 250 μm.

Figure 5. The MIMETAS OrganoReady® Colon Organoid Platform Uses IntestiCult™ to Create an Advanced Physiologically Relevant Model for Gastrointestinal Toxicity Testing and Barrier Integrity

(A) The OrganoReady® plate highlighting the microfluidic compartments.

(B) Schematic of the OrganoReady® microfluidic compartments where columns 1, 2, and 3 house the medium, a collagen-1 matrix, and the colon organoid tubule, respectively.

(C) Immunofluorescence staining of the colon organoid tubule confirms an adult tissue phenotype with the presence of goblet cells (Muc2), enterocytes (Occludin), and stem cells (Sox9). The 3D-lumenized structure provides apical (Ezrin) and basolateral (Integrin-β4) access to the polarized epithelium. Additionally, the organoid tubules show polarized and modulatable activity of expression of P-glycoprotein (Pgp).

(D) The OrganoReady® Colon Organoid platform supports toxicity testing, as demonstrated by dose-dependent measurements of TEER, LDH, and ATP following exposure to Afatinib (n = 4, N = 2). After 72 hrs of exposure, a dose dependent decrease in TEER, cytotoxicity, and cell viability was observed. For more information, please visit mimetas.com/en/organoready-organoid/.