Transcription factor-mediated differentiation of human iPSCs into neurons
Open accessAbstract
Accurate modeling of human neuronal cell biology has been a long‐standing challenge. However, methods to differentiate human induced pluripotent stem cells (iPSCs) to neurons have recently provided experimentally tractable cell models. Numerous methods that use small molecules to direct iPSCs into neuronal lineages have arisen in recent years. Unfortunately, these methods entail numerous challenges, including poor efficiency, variable cell type heterogeneity, and lengthy, expensive differentiation procedures. We recently developed a new method to generate stable transgenic lines of human iPSCs with doxycycline‐inducible transcription factors at safe‐harbor loci. Using a simple two‐step protocol, these lines can be inducibly differentiated into either cortical (i 3 Neurons) or lower motor neurons (i 3 LMN) in a rapid, efficient, and scalable manner (Wang et al., 2017). In this manuscript, we describe a set of protocols to assist investigators in the culture and genetic engineering of iPSC lines to enable transcription factor–mediated differentiation of iPSCs into i 3 Neurons or i 3 LMNs, and we present neuronal culture conditions for various experimental applications. © 2018 by John Wiley & Sons, Inc.
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Transcription factor driven differentiation protocols demonstrate greater reproducibility, purity, and decreased cost/generation time.
“Whilst protocols using transcription factor driven differentiation can overcome some of these challenges, demonstrating greater reproducibility, purity, and decreased cost/generation time12–14”
From Examining iPSC derived motor neuron variability and genome stability monitoring as a solution (2025) · cites this paper as [12]
The Commentary’s Background Information says integrated transcription factors enable uniformly efficient differentiation and that TF-based methods offer higher purity and a shorter timeline than small-molecule methods. The Introduction describes small-molecule methods as variable and expensive, but our copy does not establish that TF-based differentiation costs less. No page numbers are available in the indexed passages. (LLM, conf=0.84)
Experimental — accuracy not yet measuredThe i3Neuron platform yields large numbers of highly uniform neurons, which is necessary for robust population-based screens.
“which yields large quantities of highly homogeneous neurons, a prerequisite for robust population-based screens.”
From CRISPR Interference-Based Platform for Multimodal Genetic Screens in Human iPSC-Derived Neurons (2019) · cites this paper as Fernandopulle et al., 2018
Ngn2-driven generation yields glutamatergic neurons.
“In addition to the Ngn2-driven generation of glutamatergic neurons (Fernandopulle et al., 2018; Wang et al., 2017; Zhang et al., 2013) used here”
From CRISPR Interference-Based Platform for Multimodal Genetic Screens in Human iPSC-Derived Neurons (2019) · cites this paper as Fernandopulle et al., 2018
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- Authors
- Fernandopulle, M. S., Prestil, R., Grunseich, C., Wang, C., Gan, L., & Ward, M. E.
- Affiliations
- National Institutes of Health (US)
- Gladstone Institutes (US)
- Venue
- Current Protocols in Cell Biology
- Publisher
- Wiley
- Published
- 2018-05-18
- Volume
- 79
- Issue
- 1
- Pages
- e51-e51
- ISSN
- 1934-2500 (ISSN-L)
- 1934-2616
- PMC
- PMC6993937
- OpenAlex
- W2803819386
- Cited by
- 461
- References
- 41
- Citations by year
- 2026: 83 · 2025: 101 · 2024: 108 · 2023: 63 · 2022: 43 · 2021: 38 · 2020: 15 · 2019: 10
- FWCI
- 5.60
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Current Protocols in Cell Biology (publishedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- Funders
- National Institutes of Health (Z99 NS999999)
- National Institute on Aging (F30AG060722)
- National Institute of Neurological Disorders and Stroke (ZIANS003155)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Pluripotent Stem Cells Research
- Life Sciences › Neuroscience › Developmental Neuroscience › Neurogenesis and neuroplasticity mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Keywords
- induced pluripotent stem cells, neuronal differentiation, transcription factors, doxycycline-inducible expression, cortical neurons, lower motor neurons, small molecule differentiation
- MeSH terms
- Cell Differentiation, Cell Separation, Cells, Cultured, Chromatography, Affinity, Edetic Acid, Freezing, Humans, Lipids, Motor Neurons, Transcription Factors, Transfection, Reproducibility of Results, Cell Culture Techniques, Transgenes, Induced Pluripotent Stem Cells, Genotyping Techniques
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