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LIVER Publication-derived

HCC / CCA / CHC Tumoroids (Broutier/Clevers)

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

👤 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 ⏱ 28 days 📋 5 phases 🧫 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

Tissue Preparation and Dissociation Day 0–1
  1. 1 Obtain and prepare primary liver cancer tissue specimen
  2. 2 Mince tumor tissue
  3. 3 Enzymatic tissue digestion with extended incubation
  4. 4 Filter and wash digested cell suspension
Initial Culture in Isolation Media Day 1–21
  1. 5 Embed cells in basement membrane extract
  2. 6 Establish dual culture conditions: Classical and tumoroid-specific isolation media
  3. 7 Change isolation medium twice per week
  4. 8 Monitor and hand-pick contaminating healthy organoids (if needed)
First Passage and Transfer to Expansion Medium Day 14–28
  1. 9 Mechanically dissociate organoids into small fragments
  2. 10 Transfer mechanically dissociated fragments to expansion medium
  3. 11 Establish routine expansion schedule
Long-Term Culture and Maintenance Day 28 onwards (months to ~1 year)
  1. 12 Maintain organoid cultures in expansion medium
  2. 13 Test for mycoplasma monthly
  3. 14 Prepare and store frozen stocks
Characterization and Quality Control Throughout culture (days 14+)
  1. 15 Perform histological analysis
  2. 16 Perform immunohistochemistry for subtype-specific markers
  3. 17 Perform karyotype analysis

Full SOP

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Attribution

This SOP was authored by Organthis based on the published method in Broutier et al.. The originating laboratory holds no rights in this SOP and has not endorsed it unless marked Verified.

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