Overview

Phenotypic assays using various neuronal cell models are robust tools to discover new disease targets, perform HT screens, understand mechanism of action, and develop clinically relevant biomarkers. Assays performed in commercially available cell lines typically do not result in clinically relevant data, but this challenge is addressed by using human ES cells and iPS-derived neurons from patient samples. At Charles River, we have developed robust and reproducible in vitro models that are validated for multiple assay platforms and are reliable tools to screen compounds.

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    • IN VITRO MODELS
      • Cultured cells from post-mortem human brain tissue
        • Access to a tissue bank for post-mortem brain tissue samples supports the isolation of astrocytes and enriched microglia cultures to study multiple neurological diseases (e.g., Alzheimer’s disease, Parkinson’s disease)
      • Human immortalized fetal brain cells
        • The cells are derived from ventral mesencephalon and immortalized via retroviral transduction. These cells are differentiated into neurons that show dopaminergic-like phenotypes and are a good model for Parkinson’s disease.
      • Human embryonic stem cells (hESCs)
        • The pluripotency status of the ES cells is established using Oct4 staining pre- and post-differentiation. The ES cells can be differentiated into neuronal precursor cells (NPCs) and ultimately striatal neurons that are tested using a panel of antibody markers and/or branched DNA assays. Neuronal precursor cell markers include Pax6, FoxG1, FoxP1, Sox1, and Nestin, while some of the markers for mature neurons include MAP2, PSD95, Synaptophysin, and DARPP32.
      • Human-induced pluripotent stem cells (iPSC)
        • iPS cells are derived from reprogramming patient or controlling somatic cells. The iPS cells are differentiated into neurons using established protocols, followed by testing using a panel of neuronal specific markers.
      • Rodent primary neurons derived from established in vivo disease models
        • Established neuronal cells from various brain compartments validated using morphology and functional readouts across multiple diseases and readouts, including neuroinflammation and neuroregeneration.
    • IN VITRO ASSAYS

      In addition, our scientific team performs multiple assay readouts that are applicable in human and rodent cell systems:

      • High content screening using the InCell Analyzer platform (GE)
      • Electrophysiology
      • mRNA expression using branched DNA, qPCR, and in situ hybridization
      • Protein expression changes using single molecular analysis (Quanterix), HTRF® assays, MSD® assays, Western blotting, ELISA, Luminex®, and flow cytometry/FACS
      • IHC markers and histological stains with semi-quantitative image analysis and quantitative stereological analysis
      • Immunocytochemistry panels to assay cell type specific markers
      • Neuronal cell death assays
      • Phenotypic changes, including dendritic retraction
    • VALIDATION DATA

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      Figure 1: Left image – Rat midbrain cultures of dopaminergic neurons (TH, green) and nuclei (blue); Right image – Rat cortical cultures of neurons (MAP-2, red)
       

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      Figure 2: GABA expressing neurons originate from lateral ganglionic eminence
       

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      Figure 3: Dendritic retraction readout measured by increased MAP2 expression
       

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      Figure 4: High-content data showing knockdown of Tau in day 23 MSNs
       

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      Figure 5: Western blot data showing knockdown of Tau in day 23 MSNs

    Neuroscience 2017 Posters Now Available

    synapseCheck out our scientific posters from the SfN Annual Meeting.

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