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BRAIN Publication-derived

Alzheimer's Disease Cerebral Organoids with Astrocytes

Full SOP with Academic or a plan

Source Huang et al., 2022 · State Key Laboratory of Cell Biology, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences · 10.1038/s41392-022-01006-x

Authors: Shichao Huang, Zhen Zhang, Junwei Cao, Yongchun Yu, Gang Pei Duration: 60 days 5 phases Cell source: Human iPSC

Abstract

This protocol describes the generation of chimeric human cerebral organoids (chCOs) that incorporate induced astrocytes via lentiviral expression of NFIB and SOX9 transcription factors. The resulting organoids model Alzheimer's disease pathology by combining neurons and astrocytes with different APOE genotypes (APOE3 or APOE4), enabling assessment of the differential roles of astrocytic and neuronal APOE4 in tau and amyloid-beta pathology.

Cell source
Human iPSC
Application
Disease modeling

Protocol overview

17 steps across 5 phases

Phase 1 · hPSC Culture and Preparation Days -7 to 0
  1. a Maintain hPSC culture
  2. b Confirm cell viability and mycoplasma status
Phase 2 · EB Formation and Initial Organoid Setup Days 0-5
  1. a Dissociate and transduce hPSCs with lentivirus
  2. b Plate mixed cells into EB formation
  3. c Culture EBs in EB Formation Medium
Phase 3 · Organoid Induction Days 5-10
  1. a Transfer EBs to induction phase
  2. b Culture in Induction Medium
  3. c Embed organoids in Matrigel
  4. d Transfer to Expansion Medium
  5. e Transfer to Maturation Medium and orbital shaker
Phase 4 · Organoid Maturation and Assessment Days 10-60
  1. a Perform medium changes
  2. b Monitor astrocyte induction (optional milestones)
  3. c Assess neuronal maturation at day 60 (optional)
Phase 5 · Organoid Harvesting and Analysis Preparation Days 45-60
  1. a Prepare organoids for immunofluorescence staining
  2. b Embed organoids in OCT and cryosection
  3. c Prepare organoids for single-cell isolation
  4. d Prepare supernatant for biochemical assays

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Attribution

This SOP was written by Organthis based on the published method in Huang et al., 2022. Once the originating laboratory has confirmed it, the SOP is marked Verified.

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