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Glial cells
Immunocytochemistry imaging of oligodendroglial cells constitutively expressing Cas9 for gene knockouts
Flow cytometry analysis of B2M protein expression in CRISPRko-Ready Oligodendrocyte-like cells after delivery of a guide RNA (gRNA) targeting B2M.
Workflow schematics of the use of CRISPR knockout oligodendroglial cells
Immunocytochemistry staining of CRISPR knockout oligodendrocyte-like cells with O4 and MBP
Bulk RNA-seq data demonstrating equivalent results between wild-type and CRISPR knockout oligodendrocyte-like cells.
RT-qPCR for key mature oligodendrocyte genes in human iPSC-derived oligodendroglial cells
Morphology images of oligodendroglial cells constitutively expressing Cas9
Immunocytochemistry imaging of oligodendroglial cells constitutively expressing Cas9 for gene knockouts
Flow cytometry analysis of B2M protein expression in CRISPRko-Ready Oligodendrocyte-like cells after delivery of a guide RNA (gRNA) targeting B2M.
Workflow schematics of the use of CRISPR knockout oligodendroglial cells
Immunocytochemistry staining of CRISPR knockout oligodendrocyte-like cells with O4 and MBP
Bulk RNA-seq data demonstrating equivalent results between wild-type and CRISPR knockout oligodendrocyte-like cells.
RT-qPCR for key mature oligodendrocyte genes in human iPSC-derived oligodendroglial cells
Morphology images of oligodendroglial cells constitutively expressing Cas9

cat no | io1095

CRISPRko-Ready ioOligodendrocyte-like cells

Human iPSC-derived oligodendrocyte-like cells expressing Cas9

  • Cryopreserved human iPSC-derived cells powered by opti-ox, that are ready for experiments in days
  • Ideal for functional genomics, stably expressing Cas9 for gene knockouts and CRISPR screens
  • Resemble a pre-myelinating oligodendrocyte expressing MBP marker by day 8

Place your order

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Immunocytochemistry imaging of oligodendroglial cells constitutively expressing Cas9 for gene knockouts

Human iPSC-derived oligodendrocyte-like cells expressing Cas9

Flow cytometry analysis of B2M protein expression in CRISPRko-Ready Oligodendrocyte-like cells after delivery of a guide RNA (gRNA) targeting B2M.

Flow cytometry analysis reveals high knockout efficiency of beta-2 microglobulin (B2M) by lentiviral transduction

Flow cytometry analysis of B2M protein expression in CRISPRko-Ready ioOligodendrocyte-like cells after delivery of a guide RNA (gRNA) targeting B2M. gRNAs were introduced into the cells at day 2 post-thaw using lentiviral transduction and gene knockout was assessed 6 days post guide delivery. B2M was selected as a proof-of-concept target because it is widely expressed in the cells.

(A) Lentiviral transduction on day 2 with gRNA targeting B2M; approximately 52% of cells received a B2M gRNA, as measured by GFP expression. 

(B) Functionality of Cas9 upon B2M gRNA delivery is confirmed, as demonstrated by the histograms showing loss of B2M protein expression in 95% of cells (orange) compared to the cells that received a non-targeting (NT) gRNA (grey); cells gated on the GFP+ population.

Workflow schematics of the use of CRISPR knockout oligodendroglial cells

Go from seeding to knockout to readout in days

Immunocytochemistry staining of CRISPR knockout oligodendrocyte-like cells with O4 and MBP

CRISPRko-Ready ioOligodendrocyte-like cells express typical oligodendroglial-specific markers

Immunofluorescent staining of the cells at day 1 (upper panel) and day 8 (lower panel) post-revival. Marker expression is highly similar between CRISPRko-Ready and wild-type ioOligodendrocyte-like cells. At day 1, the cells are positive for the oligodendrocyte-specific marker O4 (red), and the DAPI counterstain (blue). At day 8, cells show an increased complexity and are positive for O4 (red), the myelin basic protein (MBP) (green), and the DAPI counterstain (blue). Scale bar: 100 μm.

View a step-by-step immunofluorescent staining protocol

Bulk RNA-seq data demonstrating equivalent results between wild-type and CRISPR knockout oligodendrocyte-like cells.

Whole transcriptome analysis demonstrates equivalent expression profiles between CRISPRko-Ready and wild-type ioOligodendrocyte-like cells

Bulk RNA-sequencing analysis was performed on CRISPRko-Ready and wild-type ioOligodendrocyte-like cells at day 1 and day 8. Principal component analysis (PCA) represents the variance in gene expression between the CRISPRko-Ready and wild-type ioOligodendrocyte-like cells. Differential expression analysis reveals no statistically significant differentially expressed (DE) genes between the two products, based on a log fold change threshold of 0.5 and an s-value cutoff of 0.01.

This analysis shows clustering of both products, demonstrating equivalent expression profiles at each given timepoint. Colours represent different samples; shapes represent different timepoints; n=3 replicates.

Expression levels for specific genes of interest can be requested by contacting our team at technical@bit.bio.

RT-qPCR for key mature oligodendrocyte genes in human iPSC-derived oligodendroglial cells

Key oligodendroglial genes are expressed by CRISPRko-Ready and wild-type ioOligodendrocyte-like cells

Gene expression analysis demonstrates that CRISPRko-Ready ioOligodendrocyte-like cells (CRISPRko-Ready) and wild-type ioOligodendrocyte-like cells (wild-type) have similar marker expression at two timepoints.

Following deterministic programming, the cells downregulate expression of the pluripotency genes OCT4, whilst demonstrating robust expression of relevant oligodendroglial markers, including PDGFRA, PLP1, MBP, and MAG. Gene expression levels assessed by RT-qPCR, data expressed relative to the reference (housekeeping) gene, HMBS. Data represents day 1 and day 8 post-thaw samples; cDNA samples of the parental human iPSC line (iPSC) were included as reference; n=3 technical replicates.

Morphology images of oligodendroglial cells constitutively expressing Cas9

Cells rapidly mature, transitioning into an oligodendrocyte-like morphology within 8 days

Upon deterministic programming, cells show rapid morphological changes, acquiring an OPC-like morphology by day 1 post-revival. By day 8, cells have matured and display an oligodendrocyte-like morphology. Morphology changes are highly similar between CRISPRko-Ready and wild-type ioOligodendrocyte-like cells. Brightfield images show day 1 and day 8 post-thawing; scale bar: 100 μm.

A maximum number of 20 vials applies. If you would like to order more than 20 vials, please contact us at orders@bit.bio.

Human iPSC-derived oligodendrocyte-like cells expressing Cas9 for rapid gene knockout generation and CRISPR screens

CRISPRko–Ready ioOligodendrocyte-like cells are deterministically programmed using opti‑ox™ technology to resemble a pre-myelinating oligodendrocyte state.

Constitutively expressing Cas9 nuclease, the cells are ready for guide RNA (gRNA) delivery from day 2 post-thaw. Using our optimised lentivirus gRNA delivery protocol, scientists can perform gene knockouts, pooled or arrayed CRISPR screens and start measuring readouts within a few days.

These ready-to-use O4⁺ cells remove the need to engineer Cas9-stable iPSC lines or optimise differentiation protocols, significantly reducing experimental timelines and allowing for reliable and reproducible experimental results by simply introducing gRNAs targeting the gene of interest.

These cells are a powerful tool for functional genomics, drug target identification and translational research for neurodegenerative and demyelinating diseases, such as multiple sclerosis.

Benchtop benefits

High knockout efficiency in CRISPR knockout oligodendrocyte-like cells

High knockout efficiency

Optimised protocol for lentivirus-based guide RNA delivery ensures maximal knockout efficiency.

CRISPR knockout Oligodendrocyte-like cells are highly defined and characterised

Defined

Human O4+ cells constitutively expressing Cas9, initially display a typical OPC-like morphology, and mature towards oligodendrocyte-like cells that have multiple branched processes.

O4-positive cells, oligodendroglial cells, ready from day 1 for experiments.

Quick

Cells are ready from day 2 post-thaw for gRNA delivery and rapidly mature into O4+ MBP+ oligodendrocyte-like cells.

Schematic overview of the timeline in the user manual

 

CRISPR knockout oligodendrocyte-like cells overview of the user manual timeline and gRNA delivery.

CRISPRko-Ready ioOligodendrocyte-like cells are delivered in a cryopreserved format and are programmed to rapidly mature upon revival in the recommended media. The protocol for the generation of these cells is a two-phase process: Induction which is carried out at bit.bio, Stabilisation for 1 day (Phase 1), and Maturation from day 1 to day 8 (Phase 2). Lentiviral gRNA delivery is recommended on day 2 post-thaw, and experimental readouts can be assessed by day 8 (day 6 post-gRNA delivery).

Download the user manual

Go from seeding to knockout to readout in days

 

Workflow schematics of the use of CRISPR knockout oligodendroglial cells

Product specifications

Starting material

Human iPSC line

Karyotype

46, XY*

Seeding compatibility

6, 12 and 24 well plates

Shipping info

Dry ice

Donor

Caucasian adult male (skin fibroblast),
Genotype APOE 3/4

Vial size

Small: >1 x 10⁶ viable cells, Evaluation pack**: 3 small vials of >1 x 10⁶ viable cells

Quality control

Sterility, protein expression (ICC), gene expression (RT-qPCR) and Cas9 functional validation (flow cytometry)

Differentiation method

opti-ox deterministic cell programming

Recommended seeding density

27,000 cells/cm²

User storage

LN2 or -150°C

Format

Cryopreserved cells

Product use

ioCells are for research use only

Applications

Single gene knockouts
Combinatorial gene knockouts
Pooled CRISPR screens
Arrayed CRISPR screens
High throughput screening

*High-resolution Optical Genome Mapping (OGM) has identified a ∼35mb gain of the short arm of chromosome 12

** Evaluation packs are intended for first-time users, or for existing users testing a new cell type or derivative. A user can request multiple evaluation packs as long as each one is for a different product, with only one pack allowed per product.
Technical data

Ready to generate gene knockouts

Flow cytometry analysis reveals high knockout efficiency of beta-2 microglobulin (B2M) by lentiviral transduction

Flow cytometry analysis of B2M protein expression in CRISPRko-Ready Oligodendrocyte-like cells after delivery of a guide RNA (gRNA) targeting B2M.

Flow cytometry analysis of B2M protein expression in CRISPRko-Ready ioOligodendrocyte-like cells after delivery of a guide RNA (gRNA) targeting B2M. gRNAs were introduced into the cells at day 2 post-thaw using lentiviral transduction and gene knockout was assessed 6 days post-guide delivery. B2M was selected as a proof-of-concept target because it is widely expressed in the cells.

(A) Lentiviral transduction on day 2 with gRNA targeting B2M; approximately 52% of cells received a B2M gRNA, as measured by GFP expression. 

(B) Functionality of Cas9 upon B2M gRNA delivery is confirmed, as demonstrated by the histograms showing loss of B2M protein expression in 95% of cells (orange) compared to the cells that received a non-targeting (NT) gRNA (grey); cells gated on the GFP+ population.

Highly characterised and defined

CRISPRko-Ready ioOligodendrocyte-like cells express typical oligodendroglial-specific markers

Immunocytochemistry staining of CRISPR knockout oligodendrocyte-like cells with O4 and MBP

Immunofluorescent staining of the cells at day 1 (upper panel) and day 8 (lower panel) post-revival. Marker expression is highly similar between CRISPRko-Ready and wild-type ioOligodendrocyte-like cells. At day 1, the cells are positive for the oligodendrocyte-specific marker O4 (red), and the DAPI counterstain (blue). At day 8, cells show an increased complexity and are positive for O4 (red), the myelin basic protein (MBP) (green), and the DAPI counterstain (blue). Scale bar: 100 μm.

View a step-by-step immunofluorescent staining protocol

Cells rapidly mature, transitioning into an oligodendrocyte-like morphology within 8 days

Morphology images of oligodendroglial cells constitutively expressing Cas9

Upon deterministic programming, cells show rapid morphological changes, acquiring an OPC-like morphology by day 1 post-revival. By day 8, cells have matured and display an oligodendrocyte-like morphology. Morphology changes are highly similar between CRISPRko-Ready and wild-type ioOligodendrocyte-like cells. Brightfield images show day 1 and day 8 post-thawing; scale bar: 100 μm.

Key oligodendroglial genes are expressed by CRISPRko-Ready and wild-type ioOligodendrocyte-like cells

RT-qPCR for key mature oligodendrocyte genes in human iPSC-derived oligodendroglial cells

Gene expression analysis demonstrates that CRISPRko-Ready ioOligodendrocyte-like cells (CRISPRko-Ready) and wild-type ioOligodendrocyte-like cells (wild-type) have similar marker expression at two timepoints.

Following deterministic programming, the cells downregulate expression of the pluripotency genes OCT4, whilst demonstrating robust expression of relevant oligodendroglial markers, including PDGFRA, PLP1, MBP, and MAG. Gene expression levels assessed by RT-qPCR, data expressed relative to the reference (housekeeping) gene, HMBS. Data represents day 1 and day 8 post-thaw samples; cDNA samples of the parental human iPSC line (iPSC) were included as reference; n=3 technical replicates.

Whole transcriptome analysis demonstrates equivalent expression profiles between CRISPRko-Ready and wild-type ioOligodendrocyte-like cells

Bulk RNA-seq data demonstrating equivalent results between wild-type and CRISPR knockout oligodendrocyte-like cells.

Bulk RNA-sequencing analysis was performed on CRISPRko-Ready and wild-type ioOligodendrocyte-like cells at day 1 and day 8. Principal component analysis (PCA) represents the variance in gene expression between the CRISPRko-Ready and wild-type ioOligodendrocyte-like cells. Differential expression analysis reveals no statistically significant differentially expressed (DE) genes between the two products, based on a log fold change threshold of 0.5 and an s-value cutoff of 0.01.

This analysis shows clustering of both products, demonstrating equivalent expression profiles at each given timepoint. Colours represent different samples; shapes represent different timepoints; n=3 replicates.

Expression levels for specific genes of interest can be requested by contacting our team at technical@bit.bio.

Product resources

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How to culture ioOligodendrocyte-like cells Video tutorial
How to culture ioOligodendrocyte-like cells
Prachi Bhagwatwar​​​​ | ​Research Assistant | bit.bio
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CRISPRko-Ready ioOligodendrocyte-like cells User manual
CRISPRko-Ready ioOligodendrocyte-like cells
V1
2025
bit.bio
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Human iPSC-Based Models of Glial Cells for Studying Neurodegenerative Disease Webinar
Human iPSC-Based Models of Glial Cells for Studying Neurodegenerative Disease
Valentina Fossati, PhD | Senior Research Investigator | The New York Stem Cell Foundation

Inês Ferreira | Senior Product Manager | bit.bio
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ioOligodendrocyte-like cells Brochure
ioOligodendrocyte-like cells
bit.bio
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ioOligodendrocyte-like cells | User Manual User manual
ioOligodendrocyte-like cells | User Manual

V2

bit.bio

2024

Download
Rethinking Developmental Biology With Cellular Reprogramming Webinar
Rethinking Developmental Biology With Cellular Reprogramming

Mark Kotter | CEO and founder | bit.bio

Marius Wernig | Professor Departments of Pathology and Chemical and Systems Biology |  Stanford University

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Precision Cellular Reprogramming for Scalable and Consistent Human Neurodegenerative Disease Models Talk
Precision Cellular Reprogramming for Scalable and Consistent Human Neurodegenerative Disease Models

Madeleine Garrett | Field Application Specialist | bit.bio

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Alzheimer’s Disease Pathogenesis: Emerging Role of Microglia Webinar
Alzheimer’s Disease Pathogenesis: Emerging Role of Microglia

Dr Matthias Pawlowski | Head, Dementia-Sensitive Hospital | University of Münster

Dr Malathi Raman | Senior Product Manager | bit.bio

Watch now
Cellular reprogramming to enable the precise and scalable manufacturing of human cells for therapeutic applications Talk
Cellular reprogramming to enable the precise and scalable manufacturing of human cells for therapeutic applications

Dr Alex Davenport | Senior Scientist | bit.bio

Talk at ELRIG Cell & Gene Therapy

2021

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Thinking of running a CRISPR-screen?
Thinking of running a CRISPR-screen?
Talk to us today about our custom screening services in all our CRISPR-Ready ioCells
Expand your research
Thinking of running a CRISPR-screen?
Talk to us today about our custom screening services in all our CRISPR-Ready ioCells
Thinking of running a CRISPR-screen?

Running CRISPR screens can be resource-intensive and require a lot of expertise and preparation. Contact us today to arrange a consultation with our screening experts.


CRISPR-Ready ioCells allow you to identify targets and screen novel compounds using physiologically relevant human cells. 

Multi cellular model for neuroinflammation studies with neurons and glia.
Study neuroinflammation in complex models
Assess cell damage, microglia activation and cytokine release
Expand your research
Study neuroinflammation in complex models
Assess cell damage, microglia activation and cytokine release
Multi cellular model for neuroinflammation studies with neurons and glia.

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Move into a multi-cellular model using different cell types:

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Tailor experimental conditions to answer your scientific question by including ioDisease Model Cells with disease-related mutations.

Modeling disease with your mutation of interest in comparison with isogenic control in oligodendrocyte-like cells
Study disease-related phenotypes
Make true comparisons with a genetically-matched control
Expand your research
Study disease-related phenotypes
Make true comparisons with a genetically-matched control
Modeling disease with your mutation of interest in comparison with isogenic control in oligodendrocyte-like cells

Build your custom disease model into the ioOligodendrocyte-like cells wild type. Use the genetically matched control in your experiments to make true comparisons in your data, being confident that even subtle variances in your data are attributable to your mutation of interest.

Start the conversation today

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Build a co-culture model
Study MBP expression in co-culture with ioGlutamatergic Neurons
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Build a co-culture model
Study MBP expression in co-culture with ioGlutamatergic Neurons
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Developing a myelination model in vitro?

Include ioGlutamatergic Neurons wild type or tailor experimental conditions to answer your scientific question by including ioDisease Model Cells with disease-related mutations, for example,

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Human iPSC-derived cells

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Consistent. Defined. Scalable.

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