Skip to content
← Back to browse
BRAIN Publication-derived

Patient-derived tumor organoids from malignant brain tumors for precision drug testing

Source Chen et al., 2022 · China Medical University Hospital, Taichung, Taiwan · 10.1038/s41598-022-20487-y

👤 Chun-Chung Chen, Hong-Wei Li, Yuan-Liang Wang, Chuan-Chun Lee, Yi-Chun Shen, Ching-Yun Hsieh, Hung-Lin Lin, Xian-Xiu Chen, Der-Yang Cho, Ching-Liang Hsieh, Jeng-Hung Guo, Sung-Tai Wei, John Wang, Shao-Chun Wang ⏱ 28 days 📋 6 phases 🧫 Patient-derived surgical tumor tissue (glioma, meningioma, lymphoma, and brain metastases)

Abstract

This protocol describes the generation and culture of patient-derived organoids (PDOs) from surgically resected malignant brain tumor tissues, including primary gliomas, meningiomas, lymphomas, and brain metastases. Tumor tissues are dissociated and cultured in serum-free medium on ultra-low attachment plates with orbital agitation, producing three-dimensional organoids within 2 weeks that recapitulate tumor histopathology and can be used for functional drug-response screening to guide personalized therapeutic decisions.

Cell source
Patient-derived surgical tumor tissue (glioma, meningioma, lymphoma, and brain metastases)
Application
Disease modeling and drug screening for personalized precision treatment

Protocol overview

31 steps across 6 phases

Tumor tissue collection and processing Day 0
  1. 1 Obtain surgical tumor specimens
PDO culture initiation and growth phase Days 1–14
  1. 1 Prepare ultra-low attachment culture plates with PDO medium
  2. 2 Seed tumor tissue pieces into culture plates
  3. 3 Set up orbital shaker in incubator
  4. 4 Perform first medium change
  5. 5 Continue medium changes every 48 hours
  6. 6 Monitor organoid development
PDO characterization by histology and immunohistochemistry Days 14–16
  1. 1 Collect organoid samples for H&E staining
  2. 2 Embed organoids in paraffin
  3. 3 Cut thin tissue sections
  4. 4 Perform hematoxylin and eosin (H&E) staining
  5. 5 Perform immunohistochemistry (IHC) for GFAP
  6. 6 Examine and compare histopathological features
PDO propagation and passaging Days 14 onwards (as needed for drug testing)
  1. 1 Assess organoid size and viability
  2. 2 Dissect organoids into smaller pieces
  3. 3 Transfer dissected organoid pieces to fresh medium
  4. 4 Continue culture maintenance
BrdU incorporation assay for drug response assessment Days 21–28 (or as needed for drug testing)
  1. 1 Prepare organoids for drug testing
  2. 2 Add test drug to organoid cultures
  3. 3 Add BrdU labeling reagent
  4. 4 Centrifuge and dry microplate
  5. 5 Fix organoids in FixDenat solution
  6. 6 Incubate with anti-BrdU-POD antibody
  7. 7 Wash wells and add substrate
  8. 8 Terminate reaction and measure absorbance
  9. 9 Normalize BrdU data by organoid diameter
Cell viability and death detection by propidium iodide and Hoechst staining Days 21–28 (as needed to confirm drug-induced cell death)
  1. 1 Treat organoids with drug and incubate
  2. 2 Wash organoids with PBS
  3. 3 Stain with Hoechst 33342 and propidium iodide
  4. 4 Image organoids by fluorescence microscopy
  5. 5 Quantify and analyze cell death

Full SOP

🔬

Create a free account to access this protocol

Join OrganMatch to unlock step-by-step procedures, reagent concentrations, QC checklists, and downloadable batch record templates.

Create free account

Already registered? Log in

Attribution

This SOP was authored by Organthis based on the published method in Chen et al., 2022. The originating laboratory holds no rights in this SOP and has not endorsed it unless marked Verified.

This wording is awaiting legal review.

Something wrong with this entry? Report an issue with this protocol

Need a commercial licence?
Use this protocol in your therapeutic or diagnostic pipeline.
Request Commercial Licence