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OTHER Publication-derived

Organoids for Drug Screening (2018)

Full SOP with Academic or a plan

Source Drost et al., 2018 · Princess Máxima Centre for Paediatric Oncology and Hubrecht Institute, Utrecht, Netherlands · 10.1038/s41586-018-0024-3

Authors: Jarno Drost, Hans Clevers Duration: 21 days 8 phases Cell source: Patient-Derived Tumor Tissue (multiple cancer types)

Abstract

This protocol describes the establishment of long-term organoid cultures from primary patient-derived tumor and matching healthy tissue samples. Tumor organoids are derived by embedding tissue samples in Matrigel and culturing under serum-free conditions with tissue-specific growth factors. These organoid cultures maintain genetic and phenotypic features of the original tumors and enable rapid drug screening and personalized medicine approaches.

Cell source
Patient-Derived Tumor Tissue (multiple cancer types)
Application
Disease modeling, Drug screening, Personalized medicine

Protocol overview

21 steps across 8 phases

Phase 1 · Tissue Sample Preparation and Processing Day 0
  1. a Obtain primary tumor and matched normal tissue biopsy samples
  2. b Mechanically dissociate tissue samples
Phase 2 · Organoid Initiation in Matrigel Day 0-1
  1. a Prepare Matrigel-containing culture wells
  2. b Apply selective culture conditions if needed
Phase 3 · Standard Culture Medium Supplementation Day 1 onwards
  1. a Prepare serum-free culture medium
  2. b Add p38 MAPK inhibitor and other tissue-specific factors if required
Phase 4 · Long-Term Culture Maintenance and Expansion Day 7 onwards
  1. a Replace culture medium regularly
  2. b Perform organoid passaging and re-plating
  3. c Cryopreserve organoids for biobanking
Phase 5 · Genetic and Phenotypic Characterization Upon organoid establishment and periodically during culture
  1. a Verify tumor origin with mutation analysis
  2. b Assess phenotypic features and biomarker expression
  3. c Perform proteomic and transcriptomic profiling
Phase 6 · Drug Screening on Patient-Derived Organoids Upon organoid establishment, typically day 14-21 of culture
  1. a Prepare organoid cultures for drug screening
  2. b Apply drug compounds and drug libraries
  3. c Assess drug-induced phenotypes and cell viability
  4. d Validate hepatotoxicity using healthy hepatocyte organoids
Phase 7 · Xenotransplantation Validation (Optional) Upon completion of in vitro drug screening
  1. a Transplant organoids into immunodeficient mice
  2. b Perform in vivo drug treatment studies
Phase 8 · Mutational Process Studies (Optional) Upon clonal organoid establishment
  1. a Establish clonal organoid cultures
  2. b Perform whole genome sequencing on clonal organoids
  3. c Study effects of chemotherapy or radiation on mutation profiles

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Attribution

This SOP was written by Organthis based on the published method in Drost et al., 2018. Once the originating laboratory has confirmed it, the SOP is marked Verified.

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