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LIVER Publication-derived

Biliary Cancer Tumouroids

Full SOP with Academic or a plan

Source Broutier et al. · Hubrecht Institute, Utrecht / Princess Máxima Center · 10.1038/nm.4438

Authors: Broutier L, Mastrogiovanni G, Verstegen MMA, Francies HE, Gavarró LM, Bradshaw CR, Allen GE, Arnes-Benito R, Sidow O, Lapthorne S, van der Laan LJW, Garnett MJ, Clevers H Duration: 28 days 5 phases Cell source: Patient-derived Liver Tumor Tissue (Resektion)

Abstract

Broutier und Clevers (Hubrecht) etablierten das erste Protokoll für Organoid-Kulturen aller drei primären Leberkrebstypen (HCC, CCA, CHC) aus Patientenresektionen. Tumoroids bewahrten histologische Architektur, genomische Landschaft und metastatisches Potenzial des Ursprungstumors auch nach langer Kulturdauer. 30 Tumor-Biomarker wurden identifiziert. Schlüsselmodifikation: Tumoroid-Medium ohne Wnt-Aktivatoren. Globaler Goldstandard für Leber-Tumoroid-Protokolle.

Cell source
Patient-derived Liver Tumor Tissue (Resektion)
Application
Liver Cancer Drug Screening

Protocol overview

17 steps across 5 phases

Phase 1 · Tissue Preparation and Dissociation Day 0-1
  1. a Obtain and prepare primary liver cancer tissue specimen
  2. b Mince tumor tissue
  3. c Enzymatic tissue digestion with extended incubation
  4. d Filter and wash digested cell suspension
Phase 2 · Initial Culture in Isolation Media Day 1-21
  1. a Embed cells in basement membrane extract
  2. b Establish dual culture conditions: Classical and tumoroid-specific isolation media
  3. c Change isolation medium twice per week
  4. d Monitor and hand-pick contaminating healthy organoids (if needed)
Phase 3 · First Passage and Transfer to Expansion Medium Day 14-28
  1. a Mechanically dissociate organoids into small fragments
  2. b Transfer mechanically dissociated fragments to expansion medium
  3. c Establish routine expansion schedule
Phase 4 · Long-Term Culture and Maintenance Day 28 onwards (months to ~1 year)
  1. a Maintain organoid cultures in expansion medium
  2. b Test for mycoplasma monthly
  3. c Prepare and store frozen stocks
Phase 5 · Characterization and Quality Control Throughout culture (days 14+)
  1. a Perform histological analysis
  2. b Perform immunohistochemistry for subtype-specific markers
  3. c Perform karyotype analysis

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Attribution

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

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