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GASTRIC Publication-derived

Gastric Cancer Tumouroids (No. 2)

Full SOP with Academic or a plan

Source 10.7150/ijbs.10248

Duration: 7 days 7 phases Cell source: Patient-Derived Gastric Cancer Tissue

Abstract

This protocol describes the isolation of primary gastric cancer cells from patient tumour tissue via enzymatic digestion and mechanical dissection, followed by organoid culture in a defined medium supplemented with growth factors and ROCK inhibitor. Cultured organoids are exposed to chemotherapeutic agents (5-FU, oxaliplatin, irinotecan, epirubicin, docetaxel) and targeted therapeutics (trastuzumab, palbociclib, imatinib) to assess drug responses and establish patient-specific sensitivity profiles for gastric cancer.

Cell source
Patient-Derived Gastric Cancer Tissue
Application
Disease modeling and drug sensitivity testing

Protocol overview

18 steps across 7 phases

Phase 1 · Tumour tissue preparation and enzymatic digestion Day 0
  1. a Tissue cutting and washing
  2. b Enzymatic digestion of tumour tissue
  3. c Tumour patch isolation
  4. d Centrifugation of tumour patches
Phase 2 · Organoid embedding and culture initiation Day 0-1
  1. a Resuspension in Matrigel
  2. b Overlay with culture medium and ROCK inhibitor
Phase 3 · Organoid expansion and maintenance Day 1-7
  1. a Culture medium composition
Phase 4 · Chemotherapeutic drug treatment Day 2-5
  1. a Organoid seeding into multiwell plates
  2. b Treatment with chemotherapeutic agents (5-FU, oxaliplatin, irinotecan, epirubicin, docetaxel)
  3. c Treatment with targeted therapeutics
Phase 5 · Viability and response assessment Day 2-5
  1. a Presto Blue cell viability assay
  2. b Apoptosis assay by flow cytometry
  3. c EdU proliferation assay
Phase 6 · Immunohistochemistry and microscopy Day 5-7
  1. a Organoid fixation and embedding
  2. b Immunohistochemical staining
  3. c Imaging
Phase 7 · Protein analysis by Western blot Day 5-7
  1. a Protein extraction and quantification
  2. b Western blot antibody incubation

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Attribution

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

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