cat no | io1007
ioGlutamatergic Neurons GBA null/R159W are opti‑ox deterministically programmed glutamatergic neurons carrying a genetically engineered compound heterozygous mutation in the GBA gene encoding the glucocerebrosidase (GCase) enzyme. These cells offer a rapidly maturing, human cell model to investigate modulation of GCase expression.
Related Parkinson's disease model cells are available with PINK1 Q456X, PRKN R275W and SNCA A53T mutations, and all can be used alongside their genetically matched control, ioGlutamatergic Neurons.
Confidently investigate your phenotype of interest across multiple clones with our disease model clone panel. Detailed characterisation data (below) and bulk RNA sequencing data (upon request) help you select specific clones if required.
per vial
A maximum number of 20 vials applies. If you would like to order more than 20 vials, please contact us at orders@ÎÞÓǶÌÊÓƵ.
Make True Comparisons
Pair the ioDisease Model Cells with the genetically matched wild-type ioGlutamatergic Neurons to investigate the impact of the GBA mutation on molecular mechanisms and cell function.
Scalable
With opti-ox technology, we can make billions of consistently programmed cells, surpassing the demands of industrial workflows.
Quick
The disease model cells and isogenic control are experiment ready as early as 2 days post revival, and form structural neuronal networks at 11 days.
ioGlutamatergic Neurons GBA null/R159W express neuron-specific markers comparably to the isogenic control
Immunofluorescent staining on post-revival day 11 demonstrates similar homogenous expression of pan-neuronal proteins TUBB3 and MAP2 (upper panel) and glutamatergic neuron-specific transporter VGLUT2 (lower panel) in ioGlutamatergic Neurons GBA null/R159W compared to the isogenic control. 100X magnification.
ioGlutamatergic Neurons GBA null/R159W form structural neuronal networks by day 11
ioGlutamatergic Neurons GBA null/R159W mature rapidly and form structural neuronal networks over 11 days, when compared to the isogenic control. Day 1 to 11 post thawing; 100X magnification.
ioGlutamatergic Neurons GBA null/R159W demonstrate gene expression of neuronal and glutamatergic-specific markers following deterministic programming
Gene expression analysis demonstrates that ioGlutamatergic Neurons GBA null/R159W and the isogenic control (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 normalised to HMBS; cDNA samples of the parental human iPSC line (hiPSC) were included as reference). Data represents day 11 post-revival samples, n=2 replicates.
Disease-related GBA is expressed in ioGlutamatergic Neurons GBA null/R159W following deterministic programming
Gene expression analysis demonstrates that ioGlutamatergic Neurons GBA null/R159W and the isogenic control (WT Control) express the GBA gene encoding the glucocerebrosidase protein. Gene expression levels were assessed by RT-qPCR (data normalised to HMBS, cDNA samples of the parental human iPSC line (hiPSC) were included as reference). Data represents day 11 post-revival samples, n=2 replicates.
GBA protein is present in ioGlutamatergic Neurons GBA null/R159W at a lower level than the wild type control
A Western blot experiment confirmed the presence of the GBA protein in ioGlutamatergic Neurons GBA null/R159W at a lower level than in the wild-type ioGlutamatergic Neurons. Day 11 cell lysates were subjected to Western blotting (20 µg protein in 40 µl per lane) using 4-20% mini protean TGX stain-free gels. Proteins were transferred onto PVDF membranes using the Trans-Blot Turbo Transfer Pack, blocked for 10 minutes, incubated with primary antibodies (GBA Invitrogen MA5-26589, 1:2000; GAPDH Abcam ab8245, 1:5000), washed three times, incubated with HRP-labelled secondary antibodies, washed three times and signal visualised by electrochemiluminescence.
1= ioGlutamatergic Neurons (wild type), 2= ioGlutamatergic Neurons GBA null/R159W.
ioGlutamatergic Neurons GBA null/R159W 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.
The recommended minimum seeding density is 30,000 cells/cm2, compared to up to 250,000 cells/cm2 for other similar commercially available products. 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 plates. This means every vial goes further, enabling more experimental conditions and more repeats, resulting in more confidence in the data.
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 (long amplicon sequencing)
Differentiation method
opti-ox deterministic cell programming
Recommended seeding density
30,000 cells/cm2
User storage
LN2 or -150°C
Format
Cryopreserved cells
Genetic modification
Compound heterozygous null/R159W mutation in the GBA gene*
Applications
Gaucher and Parkinson's disease research
Drug discovery and development
Disease modelling
Product use
ioCells are for research use only
*The null allele has a 1 base heterozygous deletion at position chr1:155,238,622 (GRCh38) located in coding exon 6, causing a frameshift resulting in a series of STOP codons (ENST00000574670.5). The second allele has a missense mutation, R159W (NM_000157.4(GBA1):c.475C>T (p.Arg159Trp))
ÎÞÓǶÌÊÓƵ
V11
ÎÞÓǶÌÊÓƵ
2024
Professor Deepak Srivastava
Professor of Molecular Neuroscience and Group Leader, MRC Centre for Developmental Disorders
King’s College London
Emmanouil Metzakopian | Vice President, Research and Development | ÎÞÓǶÌÊÓƵ
Javier Conde-Vancells | Director Product Management | ÎÞÓǶÌÊÓƵ
Chakraborty et al
Nature Communications
2023
Featuring ioGlutamatergic Neurons
Dr Ania Wilczynska | Head of Computational Genomics | Non-Clinical | ÎÞÓǶÌÊÓƵ
Innovation showcase talk at ISSCR
Marius Wernig MD, PhD | Stanford
Mark Kotter, MD, PhD | ÎÞÓǶÌÊÓƵ
Oosterveen, et al
ÎÞÓǶÌÊÓƵ & Charles River Laboratories
2023
Qiaojin Lin et al
The EMBO Journal
2023
Featuring opti-ox powered hiPSC-derived glutamatergic neurons with constitutive expression of Cas9
Mark Kotter | CEO and founder | ÎÞÓǶÌÊÓƵ
Marius Wernig | Professor Departments of Pathology and Chemical and Systems Biology | Stanford University
Madeleine Garrett | Field Application Specialist | ÎÞÓǶÌÊÓƵ
Ritsma, et al
Charles River Laboratories & ÎÞÓǶÌÊÓƵ
2022
Read this blog on glutamatergic neuron cell culture for our top tips on careful handling, cell plating and media changes to achieve success from the outset.
Further your disease research by pairing our wild type cells with isogenic disease models.