ioGlutamatergic Neurons

Human iPSC-derived glutamatergic neurons

io1001  |  Formerly known as ioNEURONS/glut, cat no: e001

ioGlutamatergic Neurons have been reprogrammed from human induced pluripotent stem cells (iPSC) using our precise reprogramming technology: opti-ox™¹ (optimized inducible overexpression). Human stem cells, within days, convert into consistent, mature, functional glutamatergic neurons providing a high quality human model for the study of neurological activity and disease.

ioGlutamatergic Neurons consist mainly of glutamatergic neurons (>80%) characterised by the expression of the glutamate transporter genes VGLUT1 and VGLUT2. The minor remaining fraction of the neuronal population express marker genes of cholinergic neurons. A bulk RNA-seq analysis shows that ioGlutamatergic Neurons have a rostral CNS identity and express the classical cortical marker genes FOXG1 and TBR1 (data not shown).

Ready-to-culture cells are suitable as models for research in cell-type specific biology, target validation and drug screening in pharmaceutical R&D, and toxicology testing.

¹ Pawlowski et al., Stem Cell Report 2017

Product specifications

Advantages

– Ready for experimentation within days
– Highly characterised and defined
– Early electrical signal
– Easy culturing

Applications

– Academic research
– Drug development
– Neurotoxicology
– High-throughput screening
– Single cell CRISPR Screening
– 3D bioprinting

For more information download our flyer

or email info@bit.bio

Consisity

Batch to batch reproducibility and homogeneity create a stable human model for excitatory neuronal activity and disease.

Speed

Ready for experimentation as early as 2 days post revival and form functional neuronal networks at 17 days.

Scalability

Industrial scale quantities at a price point that allows the cells to be used from research to screening scale.

Ease Of Use

Cells arrive programmed to rapidly mature upon revival. One medium required in a two-step protocol.

 
ioGlutamatergic Neurons generated by NGN2-driven reprogramming of iPSCs using opti-ox™ technology

Video capturing the rapid morphological changes from iPSCs upon induction of NGN2 expression using opti-ox™ cellular reprogramming. 7 day time course.

 
ioGlutamatergic Neurons express glutamatergic neuron-specific markers

TUBB3

TUBB3

VGLUT1

VGLUT1

MAP2

MAP2

VGLUT2

VGLUT2

Immunofluorescent staining on post-revival day 11 demonstrates homogenous expression of pan-neuronal proteins (MAP2 and TUBB3) and glutamatergic neuron-specific transporters (VGLUT1 and VGLUT2). Cells exhibit neurite outgrowth.

ioGlutamatergic Neurons after revival over the course of the first 11 days

Day1
Day4
Day7
Day11
Day 1 to 11 post-thawing; 400X magnification; scale bar: 100µm.

 
ioGlutamatergic Neurons display neuronal activity that matures over-time

neuronal activity

Examples of MaxOne high-resolution multi electrode array (MEA) recordings of ioGlutamatergic Neurons in BrainPhys™ media. The activity maps show firing rate (A), spike amplitude (B) and % of active electrodes (C). Results demonstrate a time-dependent increase of spontaneous activity during neuronal maturation from 2 to 3 weeks post-revival.

Iovino, M. et al., 2019, Charles River Laboratories.

Cells demonstrate a time-dependent increase of spontaneous bursting activity over a three-week period

bb-neurons_MEA-Bursting-1-opt

 

bb-neurons_MEA-Bursting-2-opt

 

bb-neurons_MEA-Bursting-3-opt

 

(A) The graph shows the % of active bursting electrodes for each time point. (B) An example of a spontaneous spike, taken at Day 8 post-revival (1 second sweep, 32 µV/-18 µV). (C) An example of a bursting phenotype, taken at Day 20 post-revival (1 second sweep, 16 µV/-16 µV). Cells were cultured in the bit.bio recommended open-source medium and recorded on 64-electrode MEAs.

NDimension (Science and Engineering) Ltd.

ioGlutamatergic Neurons co-cultured with rat-derived astrocytes demonstrate time-dependent increase in synchronous activity

bb-neurons_MEA-Synchronicity-1-opt

 

bb-neurons_MEA-Synchronicity-2-opt

 

bb-neurons_MEA-Synchronicity-3-opt

 

Array Wide Spike Detection Rate histograms (AWSDR – a graphical measure of synchrony) for 10-minute recordings on Day 8, 13 and 20 post-revival ioGlutamatergic Neurons in co-culture with primary rat-derived astrocytes. Results show prominent synchronicity on Day 13, exemplified by the ‘spikier’ nature of the associated AWSDR, which increases at Day 20. Cells were cultured in the bit.bio recommended open-source medium and recorded on 64-electrode MEAs.

NDimension (Science and Engineering) Ltd.

ioGlutamatergic Neurons show good suitability for high-throughput screening in 384-well format plates

ioGlutamatergic Neurons

Cytotoxicity CellTiter-Glo®️ (CTG) and TR-FRET (HTRF®️) assays for AKT serine/threonine kinase 1 (AKT) and Huntingtin (HTT) proteins were performed on ioGlutamatergic Neurons in 384-well plates treated with tool compound (cmp) at day 9 post-revival. Compound titration results in a concentration response curve for all three assays (mean±sd of 2 replicates). CTG assay on ioGlutamatergic Neurons shows an excellent average signal/ background ratio and high suitability for HTS. HTRF® assays on ioGlutamatergic Neurons show lower signals but with low variability, and could therefore also provide a suitable platform for HTS.

Iovino, M. et al., 2019, Charles River Laboratories.

 Cells arrive ready to plate

bit.bio ioGlutamatergic Neurons vials of cells
ioGlutamatergic Neurons arrive ready to plate
ioGlutamatergic 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: 1. Induction (carried out at bit.bio) 2. Stabilization for 4 days with Dox 3. Maintenance during which the neurons mature.

Cost effective and flexible

cost effective bit.bio ioGlutamatergic Neurons
ioGlutamatergic Neurons are compatible with plates ranging from 6 to 384 wells and are available in two vial sizes, tailored to suit your experimental needs with minimal waste. Recommended seeding density for ioGlutamatergic Neurons is 30,000 cells/cm2, compared to up to 250,000 cells/cm2 for other available products on the market. One Small vial can plate a minimum of 0.5 x 24-well plate, 0.75 x 96-well plate, or 1 x 384-well plate. One Large vial can plate a minimum of 1 x 24-well plate, 1.5 x 96-well plate, or 2 x 384-well plates.
 

Product specifications

Starting material
Human iPSC line

Donor
Caucasian adult male
(skin fibroblast)

Differentiation method
opti-ox™ cellular reprogramming
 

Karyotype
Normal (46, XY)

Vial size
Small: >0.75 x 106 viable cells
Large: >1.5 x 106 viable cells

Recommended seeding density
30,000 cells/cm2
 

Seeding compatibility
6 to 384 well plates

Quality control
Sterility, protein expression (IF) and gene expression (RT-qPCR)

User storage
LN2 or -150°C
 

Shipping info
Dry ice

Product use
These cells are for research use only
 


Supporting documentation

Frequently Asked Questions | Limited Use License | Statement of Use

 

 

 

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