StemoniX uses three of our solutions to validate their microBrain 3D Assay Ready Plates

The Challenge

Current models available to neuroscience researchers have limitations that hamper the development of new medicines. Complex models, such as post-mortem brains and animal models, more closely capture human brain complexity; however, post-mortem human brains are difficult to obtain and only represent the final disease stage. Moreover, animal models may not fully recapitulate the features and functionality of the human brain, making it difficult to translate such assays for clinical applications. While in vitro models based on simple cell culture systems can be easily scaled up, they lack the complex organization and structure of the human brain. Human induced pluripotent stem cell (hiPSC)-derived brain organoids are a very promising tool, but still display high variability and lack functional assays to assess phenotypes. The neuroscience field has been missing a model able to capture the uniqueness of human brain development and function in vitro in a highly homogenous fashion for developing new drugs to treat nervous system disorders.

The Solution

Scientists at StemoniX® tackled this gap by developing the microBrain® 3D platform. Using hiPSC-derived neural progenitor cells, they created highly homogenous 3D neural cortical spheroids that more closely resemble the human cortical brain. Their platform is comprised of mature neurons and astrocytes that are active and respond to neuromodulators similarly to primary neural cultures. Plus, they designed their platform in 96 and 384-well formats for use in conjunction with high-content screening instruments, such as the FLIPR Tetra® High-Throughput Cellular Screening System and the ImageXpress® Micro Confocal High-Content Imaging System (both from Molecular Devices), enabling homogenous assessment of functional human brain physiology in vitro.

ImageXpress Micro Confocal High-Content Imaging System

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SpectraMax i3x Multi-Mode Microplate Reader

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FLIPR Penta High-Throughput Cellular Screening System

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Products Used

The FLIPR^® Tetra^® High-Throughput Cellular Screening System provides an automated solution for identifying early leads in the drug discovery process and for evaluating drug efficacy and toxicity. With simultaneous pipette and read function, the system supports fast kinetic cellular assays. It can be quickly configured based on library size, detection mode, screening format, assay and target. As a fully integrated solution, the transition from assay development to lead optimization is seamless.

The ImageXpress^® Micro Confocal High-Content Imaging System helps you expand the boundaries of your research with the ultimate combination of speed, sensitivity and flexibility. Capture high quality images of whole organism, thick tissues, 3D spheroid assays, and cellular or intracellular events. Combined with MetaXpress^® High-Content Image Acquisition and Analysis Software, the ImageXpress Micro Confocal system provides a complete multi-dimensional, high-throughput screening solution to help you discover your next landmark scientific breakthrough.

The SpectraMax i3x Multi-Mode microplate reader measures spectral-based Absorbance, Fluorescence, and Luminescence with the added functionality of modular upgrades for Western Blot, Imaging, and Fast Kinetics with Injectors. The SpectraMax i3x reader allows you to unravel the mysteries of science by exploring cellular pathways and protein activation and expression in one system.

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The Results

The StemoniX microBrain 3D platform is a homogenous 3D neural spheroid system in 96 and 384-well format. Using the ImageXpress Micro Confocal system for brightfield capture and automated size measurement, highly homogenous size distribution with typical coefficients of variation of less than 4% across an individual plate have been observed. Immunofluorescence analysis using the ImageXpress microscope revealed that these 3D spheroids are composed of cortical neurons and astrocytes that display key markers of cellular maturity, such as synaptic proteins. Moreover, these spheroids presented spontaneous and synchronized calcium oscillations that are easily detected on both the FLIPR and ImageXpress systems. The functional maturity of the microBrain 3D platform was confirmed by examining a panel of neuromodulators with known mechanisms of action. Using the FLIPR system, modulation of neural activity was detected that correlated with the expected activity of these compounds, and confirmed that the spheroids possess functional glutamatergic and GABAergic circuits.

Finally, a targeted library of compounds with different potencies against the Zika Virus infection was tested in a toxicological case study. First, the SpectraMax® i3x Multi-Mode Microplate Reader (Molecular Devices) was employed to investigate cell toxicity of this compound library on the microBrain 3D platform. The cellular toxicity analysis was then complemented with functional toxicity, investigated in a high throughput fashion with the FLIPR system. Lastly, high-resolution videos of calcium oscillations following each treatment were generated using the ImageXpress confocal microscope. Altogether, integration of microBrain 3D in multiple platforms resulted in extensive characterization of the toxicological profile of a targeted library and demonstrated the feasibility of integrating this platform for in vitro investigations of complex neural phenotypes, toxicological profiles, and drug screening.

References

StemoniX microBrain 3D Assay Ready Plates as a Human iPSC-based Neural Spheroid Platform for High Throughput Drug Screening