Skip to content
Back to browse
BRAIN Publication-derived

Brain Organoids for Disease Modelling with Interneurons (2017)

Full SOP with Academic or a plan

Source Bagley et al., 2017 · Institute of Molecular Biotechnology of the Austrian Academy of Sciences (IMBA), Vienna, Austria · 10.1038/nmeth.4304

Authors: Joshua A Bagley, Daniel Reumann, Shan Bian, Julie Lévi-Strauss, Juergen A Knoblich Duration: 80 days 7 phases Cell source: Human iPSC

Abstract

This protocol describes a method for generating fused dorsal and ventral forebrain cerebral organoids from human pluripotent stem cells to model long-distance GABAergic interneuron migration within a single tissue construct. The fused organoid system recapitulates aspects of human brain development and enables functional studies of neuronal migration relevant to neurodevelopmental disorders.

Cell source
Human iPSC
Application
Disease modeling

Protocol overview

22 steps across 7 phases

Phase 1 · hPSC Maintenance and Preparation Days -14 to 0
  1. a Culture feeder-dependent hiPSCs
  2. b Maintain feeder-free H9 hESCs
  3. c Prepare hPSCs for embryoid body formation
Phase 2 · Embryoid Body (EB) Formation and Neural Induction Days 0-5
  1. a Generate embryoid bodies in suspension
  2. b Initiate neural induction with hES medium + bFGF and RI media
Phase 3 · Drug Patterning Treatment (Days 5-11) Days 5-11
  1. a Apply dorsal patterning (untreated control)
  2. b Apply ventral patterning (WNT inhibition + SHH enhancement)
  3. c Apply dorsal[CycA] patterning (enhanced dorsal identity)
Phase 4 · Organoid Embedding and Growth (Days 12-30) Days 12-30
  1. a Embed EBs in Matrigel droplets
  2. b Maintain organoids on orbital shaker
  3. c Supplement medium with Matrigel after day 40
Phase 5 · Organoid Maturation and Analysis (Days 30-80+) Days 30-80+
  1. a Perform qPCR analysis at day 30-40
  2. b Harvest organoids for cryosectioning and immunostaining
  3. c Perform immunofluorescence on tissue sections
  4. d Perform whole-mount and tile-scan imaging
Phase 6 · Time-Lapse Imaging of Cell Migration (Days 45-50) Days 45-50
  1. a Prepare organoid fusion slice cultures
  2. b Perform time-lapse confocal imaging
  3. c Analyze cell migration dynamics
Phase 7 · Pharmacological Perturbation (3-Week Slice Culture) Days 30-50 (as initiated)
  1. a Prepare organoid fusion slice cultures for drug treatment
  2. b Apply CXCR4 inhibitor treatment
  3. c Fix and immunostain treated slice cultures
  4. d Quantify migrated cell density

Full SOP

Full SOPs need an account and a plan

Create an account, then get free Academic access (for verified academics) or choose a plan. Plans include step-by-step procedures, reagent concentrations, QC checklists and batch record templates.

Create an account

Already registered? Log in

Attribution

This SOP was written by Organthis based on the published method in Bagley et al., 2017. Once the originating laboratory has confirmed it, the SOP is marked Verified.

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