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BRAIN Publication-derived

Prenatal Cocaine Exposure Brain Organoids

Full SOP with Academic or a plan

Source Lee et al., 2016 · National Institute on Drug Abuse, National Institutes of Health · 10.1038/npp.2016.156

Authors: Chun-Ting Lee, Jia Chen, Abigail A Kindberg, Raphael M Bendriem, Charles E Spivak, Melanie P Williams, Christopher T Richie, Annelie Handreck, Barbara S Mallon, Carl R Lupica, Da-Ting Lin, Brandon K Harvey, Deborah C Mash, William J Freed Duration: 66 days 5 phases Cell source: Human ESC (H1, H9, H14, CT2) and Human iPSC (NIH-i5, NIH-i7)

Abstract

This protocol describes the generation of self-organized 3D human neocortical organoids from pluripotent stem cells without extracellular scaffolding, designed to model neurodevelopmental effects of cocaine exposure. The organoids recapitulate key features of human neocortical development including neuroepithelial proliferation, neurogenesis, and neuronal migration, and are used to investigate CYP3A5-mediated mechanisms of cocaine toxicity during development.

Cell source
Human ESC (H1, H9, H14, CT2) and Human iPSC (NIH-i5, NIH-i7)
Application
Disease modeling and drug screening

Protocol overview

10 steps across 5 phases

Phase 1 · Embryoid Body Formation and Neuroectodermal Induction Day 0-4
  1. a hPSC Colony Dissociation
  2. b Embryoid Body Formation
  3. c Dual SMAD and FGF Pathway Inhibition
Phase 2 · Neuroepithelial Rosette Formation and Specification Day 4-16
  1. a Transfer to Neural Differentiation Medium
  2. b Adhesion Culture on Laminin-Coated Plates
  3. c Dorsal Cortical Rosette Isolation
Phase 3 · Neocortical Organoid Priming Day 16-24
  1. a Transition to Suspension Culture with FGF Priming
Phase 4 · Self-Organized Neocortical Organoid Development Day 24-66
  1. a Switch to Neuronal Differentiation Medium
  2. b Extended Suspension Culture for Organoid Maturation
Phase 5 · Drug Treatment (Optional: For Cocaine Exposure Studies) Day 32-44
  1. a Cocaine Exposure

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Attribution

This SOP was written by Organthis based on the published method in Lee et al., 2016. Once the originating laboratory has confirmed it, the SOP is marked Verified.

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