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LIVER Publication-derived

Primary Sclerosing Cholangitis Biliary Organoids

Full SOP with Academic or a plan

Source Moreno et al., 2024 · Mayo Clinic, Division of Gastroenterology and Hepatology, Rochester, Minnesota, USA · 10.1097/hc9.0000000000000454

Authors: Ana S. Garcia Moreno, Maria E. Guicciardi, Alexander Q. Wixom, Erik Jessen, Jingchun Yang, Sumera I. Ilyas, Jackie K. Bianchi, Filippo Pinto e Vairo, Konstantinos N. Lazaridis, Gregory J. Gores Duration: 15 days 4 phases Cell source: Patient-Derived Cholangiocytes (Primary Sclerosing Cholangitis and Non-PSC Controls)

Abstract

This protocol describes the generation and IL-17A stimulation of extrahepatic cholangiocyte organoids (ECO) derived from patients with primary sclerosing cholangitis (PSC) and non-PSC controls. The ECO are cultured from cholangiocytes obtained via endoscopic retrograde cholangiography, expanded in Matrigel-based organoid culture, and treated with recombinant human IL-17A to assess transcriptomic, secretomic, and genomic responses.

Cell source
Patient-Derived Cholangiocytes (Primary Sclerosing Cholangitis and Non-PSC Controls)
Application
Disease modeling; Immunological pathway interrogation in cholestatic liver disease

Protocol overview

18 steps across 4 phases

Phase 1 · Patient Sample Collection and Cholangiocyte Isolation Day 0
  1. a Endoscopic Retrograde Cholangiography (ERC) and Sample Acquisition
  2. b Bile Sample Preparation
  3. c Cytology Brush Processing
  4. d Cell Pellet Preparation
Phase 2 · Organoid Initiation and Expansion Days 0-14 (initial expansion)
  1. a Matrigel Embedding and Seeding
  2. b Initial Organoid Culture (First 3 Days)
  3. c Medium Transition to Expansion Medium
  4. d Organoid Passage
Phase 3 · IL-17A Stimulation and Analysis Day 14-Day 15 (experimental treatment)
  1. a IL-17A Treatment Preparation
  2. b IL-17A Treatment of ECO
  3. c Sample Collection for Transcriptomic Analysis
  4. d Supernatant Collection for Secretome Analysis
Phase 4 · Characterization by Immunofluorescence Day 7-Day 14 (parallel to expansion)
  1. a Whole-Mount Immunofluorescence Sample Preparation
  2. b Permeabilization and Blocking
  3. c Primary Antibody Staining
  4. d Washing and Secondary Antibody Staining
  5. e Nuclear Counterstaining and Mounting
  6. f Confocal Microscopy Imaging

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Attribution

This SOP was written by Organthis based on the published method in Moreno et al., 2024. Once the originating laboratory has confirmed it, the SOP is marked Verified.

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