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The method is highly selective, non-toxic, inexpensive, non-enzymatic, effective, time-saving and scalable.
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Invention Summary Human induced pluripotent stem cells (iPSCs) hold great promises for disease modeling and drug discovery. However, a rate-limiting step in iPSCs derivation is their separation from differentiated cells. It takes about 10 separations and passages for an iPSC clone to become stable, with minimal differentiation. Traditionally, at every passage, the researchers need to either manually pick the iPSCs from differentiated cells or mechanically scrape the differentiated cells from the plate. As such, for one 6-well plate, an experienced lab researcher typically spends 6-7 hours scraping the differentiated cells under a microscope. This procedure is long, laborious, tedious, and stressful. Rutgers scientists have developed a novel solution-based method that allows for highly selective and rapid separation of iPSCs from differentiated cells in tissue culture. The solution consists of inexpensive commercially available reagents and the method is non-enzymatic, non-toxic, scalable, time-saving, and much less labor-intensive. With the new method, the isolation and passage step for iPSCs on a 6-well plate can be accomplished in <30 min. After one separation step with the novel method, the re-plated iPSCs were shown to be >98% free of differentiated cells. Following a standard 8-minute incubation in the novel reagent, the iPSCs viability was >90% (trypan blue staining). Further, with incubation for an extended period up to 45 minutes and followed by re-plating and culturing for three days, the iPSCs exhibited no significant decreased viability. In a side-by-side comparison with a commercially available enzyme-free reagent advertised for dissociation and passaging of hPSCs, our novel reagent demonstrated remarkable selectivity in isolating iPSCs from differentiated cells (see image above). Market Application • Research reagent and protocol for iPSC-related basic research and industrial applications (toxicology, drug screening, diseases modeling, etc.) • Potential use in stem cell-based therapies Advantages The method is highly selective, non-toxic, inexpensive, non-enzymatic, effective, time-saving and scalable.