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BRAIN Publication-derived

Glioblastoma Cerebral Organoids for Transplantation

Full SOP with Academic or a plan

Source Zhang et al., 2021 · Department of Neurosurgery, Xiangya Hospital, Central South University · 10.1016/j.canlet.2020.12.013

Authors: Liyang Zhang, Fangkun Liu, Nathaniel Weygant, Junxia Zhang, Ping Hu, Zailong Qin, Jingxuan Yang, Quan Cheng, Fan Fan, Yu Zeng, Yongjian Tang, Yusheng Li, Anliu Tang, Fengqiong He, Jun Peng, Weihua Liao, Zhongliang Hu, Min Li, Zhixiong Liu Duration: 80 days 6 phases Cell source: Patient-Derived iPSC and Patient-Derived Glioma Cells

Abstract

A novel integrated system combining 3D ex vivo cerebral organoids derived from human iPSCs with in vivo orthotopic xenografts of patient-derived glioma cells (GPDCs) to recapitulate glioma biology, tumor heterogeneity, and chemotherapeutic response. This system enables faithful modeling of disease progression and evaluation of temozolomide sensitivity in patient-derived tumors.

Cell source
Patient-Derived iPSC and Patient-Derived Glioma Cells
Application
Disease modeling and drug screening

Protocol overview

40 steps across 6 phases

Phase 1 · Glioma Patient-Derived Tumor Cell (GPDC) Isolation and Culture Day 0–Ongoing
  1. a Tissue Collection and Mechanical Dissociation
  2. b Enzymatic Dissociation and Cell Isolation
  3. c Culture in Neurosphere Growth Medium
  4. d Passage and Maintenance
Phase 2 · Human Cerebral Organoid Culture from hiPSCs Day 0-Day 20+
  1. a hiPSC Culture and Maintenance
  2. b Embryoid Body Formation (Day 0)
  3. c Y-27632 Supplementation (Day 0-2)
  4. d Medium Replenishment (Day 0-5)
  5. e Neural Induction (Day 5)
  6. f Stationary Culture (Day 0-10)
  7. g Transfer to Matrigel (Day 10)
  8. h Cerebral Differentiation (Day 13 onwards)
  9. i Media Maintenance (Day 13+)
Phase 3 · GPDC Transplantation into Cerebral Organoids Day 20+
  1. a GPDC Preparation
  2. b Microinjection of GPDCs
  3. c Post-Injection Monitoring
  4. d Fixation and Harvest (Day 3, 5, 7 post-injection)
Phase 4 · Intracranial Xenograft Establishment Day 0-80+
  1. a GPDC Preparation for Injection
  2. b Intracranial Injection into Nude Mice
  3. c Post-Surgical Recovery
  4. d MRI Imaging (Day 8 Post-Surgery)
  5. e MRI Imaging (Day 80 Post-Surgery)
  6. f Limb Hemiplegia Monitoring
  7. g Terminal Procedures
Phase 5 · Histology and Immunofluorescence Analysis Variable (post-fixation)
  1. a Tissue Fixation
  2. b Paraffin Embedding
  3. c H&E Staining
  4. d Blocking with BSA
  5. e Primary Antibody Incubation
  6. f PBS Washing
  7. g Secondary Antibody Incubation
  8. h Final Washing and Mounting
Phase 6 · RNA Sequencing and Transcriptomic Analysis Variable (post-cell harvest)
  1. a RNA Extraction
  2. b RNA Integrity Assessment
  3. c Library Preparation
  4. d Sequencing
  5. e Read Quality Control
  6. f Read Mapping and Quantification
  7. g Differential Expression Analysis
  8. h Dimensionality Reduction and Clustering

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Attribution

This SOP was written by Organthis based on the published method in Zhang et al., 2021. Once the originating laboratory has confirmed it, the SOP is marked Verified.

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