ICC imaging of ioSensory Neurons - consistent, pure and easy to use sensory neurons for animal-free toxicology studies and chronic pain research

cat no | ioEA1024 Early Access

ioSensory Neurons

Highly pure human iPSC-derived sensory neurons with a defined nociceptor identity

ioSensory Neurons are a highly pure, easy to use and functional in vitro nociceptor model, enabling reliable chronic nociceptive pain research and therapeutics development for peripheral neuropathies.

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Benchtop benefits

pure_0

Highly Pure

>99% pure sensory neurons with a defined nociceptor identity by day 7 post-revival, as confirmed by single cell RNA sequencing.

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Functional

ioSensory Neurons display spontaneous activity and a functional nociceptor phenotype, as shown by calcium mobilisation in response to specific TRP agonists.

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Easy to use

Cryopreserved cells arrive, programmed to mature rapidly upon revival. Simple two-step protocol using one medium – no mitomycin C treatment is necessary.

Technical data

Ready within days

opti-ox precision reprogrammed ioSensory Neurons rapidly form a homogenous neuronal population

Time-lapse video capturing the rapid and homogeneous neuronal phenotype acquisition upon thawing of cryopreserved ioSensory Neurons. 14 day time course.

Highly characterised and defined

ioSensory Neurons express pan-sensory neuron-specific markers 

bitbio-ioSensory_Neurons-ICC_Comparison V2

Immunofluorescent staining of ioSensory Neurons at day 14 post-revival. The upper panel shows that ioSensory Neurons are positive for BRN3A (red), the pan-neuronal marker MAP2 (green), and the DAPI counterstain (blue), and demonstrates that all MAP2 positive neurons have a sensory neuronal identity. The lower panel shows that ioSensory Neurons are positive for ISL1 (magenta), PRPH (red), the pan-neuronal marker TUBB3 (green), and the DAPI counterstain (blue).

ioSensory Neurons show visible neuronal networks by day 7

bitbio-ioSensory_Neurons-Morphology_Timeline-correction

Upon reprogramming, rapid morphological changes are observed in the cells, with neurons identified by day 4 post-revival. Visible neuronal networks are observed by day 7 post-thaw. Images show day 1 to 14 post-thawing; 10X magnification.

Single cell RNA-sequencing shows ioSensory Neurons form a pure population (>99%) of sensory neurons

bitbio-ioSensory_Neurons-scRNA-mature-correct-key-small-V2

Single cell RNA-sequencing analysis was performed with ioSensory Neurons at four specific timepoints (days 7, 10, 14 and 17). By day 7, the population has a distinct expression profile indicating a pure population (>99%) of post-mitotic sensory 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 ioSensory Neurons express key pan-sensory neuronal markers

bitbio-ioSensory_Neurons-scRNA-nociceptor-pansensory-correction

Single cell RNA-sequencing analysis was performed with ioSensory Neurons at four specific timepoints (days 7, 10, 14 and 17). By day 7, the expression of pan-sensory neuron marker genes, ISL1, ISL2, PRPH, BRN3A, together with the pan-neuronal markers TUBB3 and MAP2, could be detected in post-mitotic sensory 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 ioSensory Neurons express key nociceptor markers

bitbio-ioSensory_Neurons-scRNA-key-nociceptor

Single cell RNA-sequencing analysis was performed with ioSensory Neurons at four specific timepoints (days 7, 10, 14 and 17). By day 7, the expression of key nociceptor marker genes (NTRK1 and TRPV1) could be detected in post-mitotic sensory neurons. 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

ioSensory Neurons display spontaneous activity that matures over time
bitbio-ioSensory_Neurons-MEA_Data

Multi electrode array (MEA) recordings of ioSensory Neurons at days 6 and 17. 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 day 6 to day 17. Analysis was performed on a Maxwell Biosystem's MaxTwo multi-well system. Note, this data is from cells in continuous culture and not cryopreserved cells.

ioSensory Neurons display a functional nociceptor phenotype
bitbio-ioSensory_Neurons-Calcium_Mobilization
Calcium mobilisation imaging upon stimulation of ioSensory Neurons with pharmacological agonists targeting thermosensitive TRP channels such as TRPV1 (capsaicin), TRPM3 (CIM0216) and TRPM8 (WS-12). Active traces represent the increase in intracellular calcium mobilisation of individual cells upon exposure to noxious stimuli but not to vehicle at day 17. This indicates that cells display a functional nociceptor phenotype within 17 days. Note, this data is from cells in continuous culture and not cryopreserved cells.
Calcium imaging of ioSensory Neurons demonstrates activity following treatment with a TRPM3 agonist
Representative video of ioSensory Neurons displaying calcium mobilisation in response to treatment with the TRPM3 agonist, CIM0216. Note, this data is from cells in continuous culture and not cryopreserved cells.

Cells arrive ready to plate

bitbio-iosensory_neurons-diagram

ioSensory 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 7 days (Phase 1), and Maintenance (Phase 2) during which the ioSensory Neurons mature. Phases 1 and 2 after revival of cells are carried out by the customer.

Product information

Starting material

Human iPSC line

Karyotype

Normal (46, XY)

Seeding compatibility

6, 12, 24, 96 and 384 well plates

Shipping info

Dry ice

Donor

Caucasian adult male (skin fibroblast)

Vial size

Small: >2 x 10⁶ viable cells

Quality control

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

Differentiation method

opti-ox cellular reprogramming

Recommended seeding density

60,000 cells/cm²

User storage

LN2 or -150°C

Format

Cryopreserved cells

Product use

ioCells are for research use only

Applications

Chronic pain research & drug development
MEA analysis
Calcium imaging
Transcriptome analysis
Neurotoxicology

Product resources

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

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

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

Watch now
Mastering Cell Identity In A Dish: The Power Of Cellular Reprogramming Webinar
Mastering Cell Identity In A Dish: The Power Of Cellular Reprogramming

Prof Roger Pedersen | Adjunct Professor and Senior Research Scientist at Stanford University 

Dr Thomas Moreau | Director of Cell Biology Research | bit.bio

Watch now
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

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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