bit_bio ioGABAergic Neurons 1-3-comp

cat no | ioEA1003S Early Access

ioGABAergic Neurons

Human iPSC-derived GABAergic neurons

ioGABAergic Neurons are human induced pluripotent stem cell (iPSC)-derived GABAergic neurons, precision reprogrammed using opti-ox™ technology. Within 12 days post-revival, ioGABAergic Neurons form a pure, defined population that is ready for experimentation, expressing (>95%) classical marker genes including GAD1, GAD2, VGAT, DLX1 and DLX2. Calcium imaging has been used to demonstrate that ioGABAergic Neurons display spontaneous activity.

ioGABAergic Neurons provide an easy to use, high-quality human model for the study of neurological circuitry, neurodevelopmental and neurodegenerative disease research and drug development. 

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Characteristics that inspire confidence

quick_0

Quick

Rapidly maturing inhibitory neurons that are ready to use within 12 days, post-revival.

defined_0

Defined

>95% of cells express key GABAergic phenotypic markers GAD1, GAD2, VGAT, DLX1 and DLX2.

functional_0

Functional

Calcium imaging demonstrates that ioGABAergic Neurons display spontaneous activity.

Product information

Starting material

Human iPSC line

Karyotype

Normal (46, XY)

Seeding compatibility

6, 12, 24 & 96 well plates

Shipping info

Dry ice

Donor

Caucasian adult male (skin fibroblast)

Vial size

Small: >3 x 10⁶ viable cells

Quality control

Sterility, protein expression (ICC) and gene expression (RT-qPCR)

Differentiation method

opti-ox cellular reprogramming

Recommended seeding density

150,000 cells/cm²

User storage

LN2 or -150°C

Format

Cryopreserved cells

Product use

ioCells are for research use only

Applications

Disease research
Co-culture studies
Calcium imaging
Transcriptome analysis
MEA analysis

Technical data

Ready within days

ioGABAergic Neurons generated by transcription factor-driven reprogramming of iPSCs using opti-ox technology

Time-lapse video capturing the rapid and homogeneous neuronal phenotype acquisition upon thawing of cryopreserved ioGABAergic Neurons. 12 day time course.

Highly characterised and defined

ioGABAergic Neurons express GABAergic neuron-specific markers

ioGABAergic Neurons ICC GABA
GABA
ioGABAergic Neurons ICC MAP2
MAP2
ioGABAergic Neurons ICC DAPI
DAPI
ioGABAergic Neurons ICC merge
MERGE

Immunofluorescent staining on day 10 post-revival demonstrates that ioGABAergic Neurons are positive for GABA (yellow), the pan-neuronal marker MAP2 (red), and the DAPI counterstain (blue).

ioGABAergic Neurons show mature neuronal networks by day 12

ioGABAergic Neurons Brightfield Day 1
DAY 1
ioGABAergic Neurons Day 3 Brightfield
DAY 3
ioGABAergic Neurons Day 7 Brightfield
DAY 7
ioGABAergic Neurons Day 12 Brightfield
DAY 12

Upon reprogramming, ioGABAergic Neurons show rapid morphological changes, with neurons identified by day 7 post-revival. Mature neuronal networks are observed by day 12. Images show day 1 to 12 post-thawing; 10X magnification; scale bar: 200 μm.

Single-cell RNA sequencing shows ioGABAergic Neurons form a pure population (>95%) of mature GABAergic neurons

ioGABAergic Neurons UMAP scRNAseq day 0-14

Single cell RNA sequencing analysis was performed with ioGABAergic Neurons at three specific timepoints (day 0, 7 and 14). By day 7, the population has a distinct expression profile indicating a pure population (>95%) of mature post-mitotic GABAergic neurons. Gene expression was assessed by 10x Genomics scRNA-sequencing. Note, this data is from cells in continuous culture and not cryopreserved cells.

Single cell RNA sequencing shows mature ioGABAergic Neurons express key GABAergic markers

UMAP GABA markers

Single cell RNA sequencing analysis was performed with ioGABAergic Neurons at three specific timepoints (day 0, 7 and 14). By day 7, the expression of key GABAergic marker genes (GAD1, GAD2, SLC32A1 (VGAT), DLX2, DLX5), together with the pan-neuronal marker MAP2, could be detected in post-mitotic GABAergic neurons. Gene expression was assessed by 10x Genomics scRNA-sequencing. Note, this data is from cells in continuous culture and not cryopreserved cells.

Single-cell RNA sequencing shows mature ioGABAergic Neurons show minimal expression of markers indicative of other neuronal lineages

UMAP scRNA-seq non-GABA markers

Single cell RNA sequencing analysis was performed with ioGABAergic Neurons at three specific timepoints (day 0, 7 and 14). By day 14, the expression of markers of non-GABAergic neuronal lineages (TH - dopaminergic, TPH1 - serotonergic, CHAT - cholinergic, SLC17A6 (VGLUT2) - glutamatergic, SLC17A7 (VGLUT1) - glutamatergic, SLC18A3 (VACHT) - cholinergic) are largely absent. Gene expression was assessed by 10x Genomics scRNA-sequencing. Note, this data is from cells in continuous culture and not cryopreserved cells.

Single cell RNA sequencing indicates that ioGABAergic Neurons are of the SST sub-type

UMAP SST subtype
Single cell RNA sequencing analysis was performed with ioGABAergic Neurons at three specific timepoints (day 0, 7 and 14). By day 7, cells appear to largely express the somatostatin (SST) marker indicating that this population is of the SST sub-type. Gene expression was assessed by 10x Genomics scRNA-sequencing. Note, this data is from cells in continuous culture and not cryopreserved cells.

Rapid gain of functional activity

Calcium imaging of ioGABAergic Neurons demonstrates spontaneous activity
Video showing spontaneous activity of ioGABAergic Neurons, subjected to calcium imaging at day 16 post-revival.

Cells arrive ready to plate

ioGABAergic neurons 12 day timeline

ioGABAergic Neurons 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 three-phase process: Induction, which is carried out at bit.bio (Phase 0), Stabilisation for 3 days (Phase 1), and Maintenance (Phase 2) during which the ioGABAergic Neurons mature. Phases 1 and 2 after revival of cells are carried out at the customer site.

Product resources

Studying Neurodevelopment with Stem Cells | Webinar Webinar
Studying Neurodevelopment with Stem Cells | Webinar

Dr Deepak Srivastava | King’s College London

Watch now
Modelling human neurodegenerative diseases in research & drug discovery Webinar
Modelling human neurodegenerative diseases in research & drug discovery

Dr Mariangela Iovino | Group Leader | Charles River
Dr Tony Oosterveen | Senior Scientist | bit.bio

Watch now

Differentiating iPSCs - which approach works best? 

Download this infographic to find out how the approach used to generate human iPSC-derived cells influences purity, batch consistency and protocol speed. 

bit.bio-opti-ox-infographic in hand-Aug 2022

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