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LUNG Publication-derived

Patient-derived lung cancer organoids as in vitro cancer models for therapeutic screening

Source Kim et al., 2019 · Asan Institute for Life Sciences, Seoul, South Korea · 10.1038/s41467-019-11867-6

👤 Minsuh Kim, Hyemin Mun, Chang Oak Sung, Eun Jeong Cho, Hye-Joon Jeon, Sung-Min Chun, Da Jung Jung, Tae Hoon Shin, Gi Seok Jeong, Dong Kwan Kim, Eun Kyung Choi, Seong-Yun Jeong, Alison M. Taylor, Sejal Jain, Matthew Meyerson, Se Jin Jang ⏱ 21 days 📋 7 phases 🧫 Patient-Derived Lung Cancer Tissues (Multiple Histological Subtypes)

Abstract

This protocol describes the establishment and culture of lung cancer organoids (LCOs) and normal bronchial organoids (NBOs) from patient-derived lung tissues. LCOs maintain the histological, genetic, and phenotypic characteristics of original tumors while enabling rapid patient-specific drug screening and therapeutic response prediction in a 3D culture system.

Cell source
Patient-Derived Lung Cancer Tissues (Multiple Histological Subtypes)
Application
Disease modeling and drug screening

Protocol overview

43 steps across 7 phases

Tissue Preparation and Quality Control Day 0-1
  1. 1 Collect and transport tissue samples
  2. 2 Wash tissue samples
  3. 3 Section tissue samples
  4. 4 Dissociate tissue and prepare cell suspension
  5. 5 Triturate and filter cell suspension
  6. 6 Centrifuge and prepare quality control sample
Lung Cancer Organoid (LCO) Culture Initiation Day 1-7
  1. 1 Prepare MBM (Minimum Basal Medium)
  2. 2 Seed LCOs in Matrigel
  3. 3 Add culture medium
  4. 4 Incubate and observe organoid formation
  5. 5 Change medium every 4 days
LCO Passaging and Expansion Day 8-21 and beyond
  1. 1 Determine passaging time point
  2. 2 Harvest organoids from Matrigel
  3. 3 Wash organoid pellet
  4. 4 Dissociate organoids to single cells
  5. 5 Inactivate trypsin and centrifuge
  6. 6 Wash and resuspend cells
  7. 7 Solidify Matrigel and add medium
  8. 8 Continue culture and monitor growth
Normal Bronchial Organoid (NBO) Culture Day 1-7 and beyond
  1. 1 Prepare MBM + WNA medium
  2. 2 Seed NBOs in Matrigel
  3. 3 Add MBM + WNA medium
  4. 4 Incubate and monitor organoid formation
  5. 5 Passage NBOs using same protocol as LCOs
Cryopreservation and Banking Passage 3–4
  1. 1 Determine organoid size for banking
  2. 2 Harvest and clean organoids
  3. 3 Prepare freezing medium
  4. 4 Resuspend organoids in freezing medium
  5. 5 Aliquot into cryogenic vials
  6. 6 Freeze organoids gradually
  7. 7 Store in gas phase nitrogen tank
Organoid Thawing and Viability Assessment As needed for experiments
  1. 1 Retrieve and thaw organoid vial
  2. 2 Transfer to culture medium
  3. 3 Centrifuge and resuspend
  4. 4 Seed thawed organoids
  5. 5 Add medium and assess viability
Drug Sensitivity Testing Passage 2+, Days 10–16
  1. 1 Prepare organoids for drug screening
  2. 2 Prepare organoid suspension for seeding
  3. 3 Allow organoid formation
  4. 4 Prepare and add drug solutions
  5. 5 Monitor organoid response
  6. 6 Measure cell viability
  7. 7 Calculate IC50 values

Full SOP

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Attribution

This SOP was authored by Organthis based on the published method in Kim et al., 2019. The originating laboratory holds no rights in this SOP and has not endorsed it unless marked Verified.

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