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cat no | io1014

ioGlutamatergic Neurons
MAPT N279K/N279K™

Human iPSC-derived FTD disease model

A rapidly maturing, consistent and scalable system to study frontotemporal dementia (FTD).

ioGlutamatergic Neurons MAPT N279K/N279K are opti-ox™ precision reprogrammed glutamatergic neurons carrying a genetically engineered homozygous N279K mutation in the MAPT gene encoding the microtubule-associated protein tau.

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Benchtop benefits

Disease related phenotype

High content ICC image analysis shows hyperphosphorylation of tau compared to the wild-type control.

Make True Comparisons

Pair the ioDisease Model Cells with the genetically matched wild-type ioGlutamatergic Neurons to directly investigate the impact of mutant tau protein on disease.

Quick

The disease model cells and genetically matched control are experiment ready as early as 2 days post revival, and form structural neuronal networks at 11 days.

Disease-related phenotype

Hyperphosphorylation of tau observed in the disease model cells compared to the wild type control

Hyperphosphorylation of tau in glutamatergic neurons FTD disease model cells carrying MAPT P301S or N279K mutation

ioGlutamatergic Neurons disease model cells carrying MAPT P301S/P301S, MAPT N279K/WT and MAPT N279K/N279K mutations show hyperphosphorylation when compared to the wild type control ioGlutamatergic Neurons (WT) at day 21. The bar graphs show total Tau, pTau217/total Tau, pTau202/5/total Tau or pTau404/total Tau in cell bodies, analysed by immunocytochemistry. Statistical analyses performed on 5 cellular replicates in the same plate. Bars showing mean, error bars showing standard deviation. Statistics calculated by one way ANOVA and Tukey posthoc analysis. Data courtesy of Charles River Laboratories.

The expression of the tau 4R isoform transcript is relevant for the study of frontotemporal dementia (FTD). Bulk RNA-sequencing data for ioGlutamatergic Neurons, the genetically matched control for the MAPT disease model cells, show an equimolar tau 3R/4R ratio at day 11, similar to the ratio in the adult human brain, making the wild type and disease model cells a suitable system for investigating FTD.

Highly characterised and defined

ioGlutamatergic Neurons MAPT N279K/N279K express neuron-specific markers comparably to the wild type control

ioGlutamatergic Neurons MAPT N279K/N279K disease model cells express neuron-specific markers comparably to wild type control

Immunofluorescent staining on post-revival day 11 demonstrates similar homogenous expression of pan-neuronal proteins MAP2 and TUBB3 (upper panel) and glutamatergic neuron-specific transporter VGLUT2 (lower panel) in ioGlutamatergic Neurons MAPT N279K/N279K compared to the genetically matched control. 100X magnification.

ioGlutamatergic Neurons MAPT N279K/N279K form structural neuronal networks by day 11

ioGlutamatergic Neurons MAPT N279K/N279K disease model cells morphology by brightfield imaging over 11 days post-revival

ioGlutamatergic Neurons MAPT N279K/N279K mature rapidly and form structural neuronal networks over 11 days, highly similar to the genetically matched control. Day 1 to 11 post thaw; 100X magnification.

ioGlutamatergic Neurons MAPT N279K/N279K demonstrate gene expression of neuronal-specific and glutamatergic-specific markers following reprogramming

ioGlutamatergic Neurons MAPT N279K/N279K key neuronal marker gene expression analysis by RT-qPCR

Gene expression analysis demonstrates that ioGlutamatergic Neurons MAPT N279K/N279K and wild-type ioGlutamatergic Neurons (WT Control) lack the expression of pluripotency markers (NANOG and OCT4) at day 11, whilst robustly expressing pan-neuronal (TUBB3 and SYP) and glutamatergic-specific (VGLUT1 and VGLUT2) markers, as well as the glutamate receptor GRIA4. Gene expression levels were assessed by RT-qPCR. Data is shown relative to the parental human iPSC control (hiPSC), normalised to HMBS. Data represents day 11 post-revival samples; n=2 biological replicates.

Disease-related MAPT is expressed in ioGlutamatergic Neurons MAPT N279K/N279K following reprogramming

ioGlutamatergic Neurons MAPT N279K/N279K expression analysis of MAPT gene by RT-qPCR

RT-qPCR analysis demonstrates a similar expression level of the MAPT gene in both wild type ioGlutamatergic Neurons (WT Control) and ioGlutamatergic Neurons MAPT N279K/N279K at day 11 post-revival (n=2 replicates). cDNA samples of the parental human iPSC line (hiPSC) were included as a reference.

Cells arrive ready to plate

ioGlutamatergic Neurons MAPT N279K/N279K arrive ready to plate

ioGlutamatergic Neurons MAPT N279K/N279K are delivered in a cryopreserved format and are programmed to mature rapidly upon revival in the recommended media. The protocol for the generation of these cells is a two-phase process: Phase 1, Stabilisation for 4 days; Phase 2, Maintenance, during which the neurons mature. Phases 1 and 2 after revival of cells are carried out by the customer.

Industry leading seeding density

Do more with every vial

ioGlutamatergic Neurons MAPT N279K/N279K seeding density in 96- and 384-well plates

ioGlutamatergic Neurons MAPT N279K/N279K cells are compatible with plates ranging from 6 to 384 wells.

The recommended seeding density is 30,000 cells/cm², compared to up to 500,000 cells/cm² for other similar products on the market.

This means scientists are able to do more with every vial and expand experimental design within budget without losing out on quality. Resulting in more experimental conditions, more repeats, and more confidence in the data.

One small vial can plate a minimum of 0.7 x 24-well plate, 1 x 96-well plate, or 1.5 x 384-well plate.

Product information

Starting material

Human iPSC line

Karyotype

Normal (46, XY)*

Seeding compatibility

6, 12, 24, 48, 96 & 384 well plates

Shipping info

Dry ice

Donor

Caucasian adult male (skin fibroblast)

Vial size

Small: >1 x 10⁶ viable cells

Quality control

Sterility, protein expression (ICC), gene expression (RT-qPCR) and genotype validation (Sanger sequencing)

Differentiation method

opti-ox cellular reprogramming

Recommended seeding density

30,000 cells/cm²

User storage

LN2 or -150°C

Format

Cryopreserved cells

Genetic modification

Homozygous N279K missense mutation in the MAPT gene

Applications

FTD research
Drug discovery and development
Disease modelling
High content imaging
Western blotting
Electrophysiological assays (MEA)
Co-culture studies

Product use

ioCells are for research use only

*ioGlutamatergic Neurons MAPT N279K/N279K disease model cells can be used with the parental wild type ioGlutamatergic Neurons (io1001) as a control. The two products are genetically matched except for a ~850Kb amplification on 20q11.21 in the disease model cells, identified by Optical Genome Mapping.

Supporting documentation

User Manual

Limited Use License & Statement of Use

Product resources

Webinar

Addressing the Reproducibility Crisis | Driving Genome-Wide Consistency in Cellular Reprogramming

Dr Ania Wilczynska | Head of Computational Genomics | Non-Clinical | bit.bio

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Industrialising Cellular Reprogramming: Leveraging opti-ox™ Technology to Manufacture Human Cells with Unprecedented Consistency

Innovation showcase talk at ISSCR

Marius Wernig MD, PhD | Stanford

Mark Kotter, MD, PhD | bit.bio

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Poster

Modelling neurodegeneration: Human isogenic system to study FTD & ALS

Dr Tony Oosterveen, et al.

bit.bio & Charles River Laboratories

2023

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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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Talk

Precision Cellular Reprogramming for Scalable and Consistent Human Neurodegenerative Disease Models

Madeleine Garrett | Field Application Specialist | bit.bio

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Poster

Development and characterisation of a robust in vitro disease model to study tauopathies

Laila Ritsma et al.

Charles River Laboratories & bit.bio

2022

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Poster

Validation of ALS-relevant phenotypes in precision reprogrammed iPSC-derived glutamatergic Neurons containing a TDP-43 M337V mutation.

Laila Ritsma et al.

Charles River Laboratories & bit.bio

2022

Download

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Read this blog for top tips on cell culture of human iPSC-derived glutamatergic neurons to help you overcome some of the most common issues. By using careful handling, it’s possible to achieve regular and consistent cell culture success!

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