RaftNotes describe applications and protocols for use with the CellRaft ® Technology
These publications cover materials and methods, tips, tricks, examples, and unique array formats that help our users achieve their best research results.
In the last decade, several protocols and commercial kits have been launched to help differentiate iPSCs into multicellular, neuronal organoids that closely resemble human brain development, including region-specific cellular composition and functional physiology. However, the adoption of these organoid models is still limited to relatively low throughout applications, as the workflows are hampered by challenges in reproducibility and scalability, as well as being manually intensive. Here we report the use of the CellRaft® Technology, to develop and enable streamlined, reproducible organoid workflows that offer reliable imaging, software-guided selection, and automated isolation of single organoids for downstream applications.
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Despite significant strides in technology and increased access for researchers, iPSC culture remains a challenge, thereby limiting its widespread utility for therapeutic and research use. Common pain points include:
1) iPSC lines are sensitive and easily perturbed, requiring constant maintenance.
2) Poor culture conditions and cell line instability can lead to spontaneous differentiation and loss of pluripotency.
3) Generating monoclonal cell lines is incredibly challenging, with low efficiency and lack of proven clonality.
4) The labor, cost, and reagent burden associated with iPSC maintenance and workflows are incredibly high and often prohibitive for the end user.
Download this RaftNote to see how some of these challenges can be overcome.
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The advancement of 3D culture systems has transformed cell-based assays for biological research and drug discovery due to their ability to re-capitulate the structure and cellular complexity of in vivo tissues. Organoids are unique due in their ability to self-organize and closely replicate in vivo pathophysiology. Most laboratory instruments employed to assess endpoints using traditional organoid cultures have limitations that preclude assessment of the heterogeneity within the population or retrieval of single, intact 3D structures for downstream applications.
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The AIR System offers unique advantages over technologies, such as flow cytometry and droplet dispensers, designed to assess cells in suspension. The limitations to other technologies include impacts on cell viability, inability to interrogate small numbers of cells, and reliance on fluorescent markers or staining for cell characterization. To demonstrate the value of the CellRaft technology for use in suspension cell line development, we have established methodologies for attaching suspension cells to the CellRaft Arrays during single cell expansion while still allowing the cells to expand in suspension after the clone of interest has been isolated.
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Therapeutic proteins play an essential role in the biological pharmaceutical market and are used in the treatment of many diseases, such as diabetes, cancer, and anemia. One of the main goals in recombinant protein development is the establishment of high-quality monoclonal cell lines that consistently express large amounts of the given protein. Chinese hamster ovary (CHO) cell lines have dominated the industry as commercial hosts for recombinant protein production; however, the process of generating a homogenous CHO cell line is not trivial.
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Gene editing workflows require transformation of a large number of cells, followed by isolation of individual cells from the larger population to establish clonal colonies. Given the large number of gene edits required for contemporary research and the labor-intensive components of the workflow, there is an unmet need to automate post-transformation cloning.
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Functional genomic analysis provides a connection between cellular phenotypes, such as drug responsiveness, and their underlying genomic precursors, such as sequence variation or differential gene expression.
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One of the benefits of the CellRaft Technology is the isolation and recovery of single cells even from low cell titer samples. This RaftNote describes a new cell seeding insert that significantly increases the number of cells available for isolation when low cell titer samples are used.
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Single cell genomic analysis has already impacted a range of life science fields, including tumor clonality, neuronal mosaicism, and drug resistance mechanisms. The CellRaft Technology uniquely enables the sorting and isolation of single nuclei for downstream analysis. Here we provide optimal methods for imaging nuclei on CellRaft Arrays, and downstream molecular biology protocols for the amplification and preparation of nucleic acid prior to sequencing.
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Sorting and isolating single cells is a key sample preparation step in many contemporary workflows including genome editing by CRISPR/Cas9, single cell genomics and differentiation of induced pluripotent stem cells (IPSCs) along with a broad range of other methods. Here, we present optimized methods for staining live cells on the Cell Microsystems CellRaft Array, a microwell array with releasable single cell isolation features called CellRafts.
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