Induced pluripotent stem cells (iPSCs) derived from human somatic cells have created new opportunities to generate disease-relevant cells. Thus, as the use of patient-derived stem cells has become more widespread, having a workflow to monitor each line is critical. This ensures iPSCs pass a suite of quality control measures, promoting reproducibility across experiments and between labs. With this in mind, we established a multistep workflow to assess our newly generated iPSCs for variations and reproducibility relative to each other and iPSCs obtained from external sources. Our benchmarks for evaluating iPSCs include examining iPSC morphology and proliferation in two different media conditions and evaluating their ability to differentiate into each of the three germ layers, with a particular focus on neurons. Genomic integrity in the human iPSCs was analyzed by G-band karyotyping and a qPCR-based test for the detection of hotspot mutations test. Cell-line identity was authenticated by Short Tandem Repeat (STR) analysis. Using standardized dual SMAD inhibition methods, all iPSC lines gave rise to neural progenitors that could subsequently be differentiated into cortical neurons. Neural differentiation was analyzed qualitatively by immunocytochemistry and quantitatively by qPCR for progenitor, neuronal, cortical, and glial markers. Taken together, we present a multistep quality control workflow to evaluate variability and reproducibility across and between iPSCs.

中文翻译：

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标准化的质量控制工作流程，用于评估人iPSC向神经元的重现性和分化潜能

源自人类体细胞的诱导性多能干细胞（iPSC）为产生与疾病相关的细胞提供了新的机会。因此，随着源自患者的干细胞的使用变得越来越普遍，具有监控每个细胞系的工作流程至关重要。这样可确保iPSC通过一套质量控制措施，从而提高整个实验之间以及实验室之间的可重复性。考虑到这一点，我们建立了一个多步骤工作流，以评估我们新生成的iPSC彼此之间以及从外部来源获得的iPSC之间的变异和可重复性。我们评估iPSC的基准包括检查iPSC的形态和在两种不同培养基条件下的增殖以及评估其分化为三个细菌层的能力，尤其是神经元。通过G带核型分析和基于qPCR的检测热点突变测试的分析，分析了人类iPSC中的基因组完整性。通过短串联重复序列（STR）分析验证了细胞系的身份。使用标准化的双重SMAD抑制方法，所有iPSC系都产生了神经祖细胞，这些神经祖细胞随后可以分化为皮层神经元。通过免疫细胞化学定性分析神经分化，并通过qPCR定量分析祖细胞，神经元，皮层和神经胶质标志物。综上所述，我们提出了一个多步骤质量控制工作流程，以评估iPSC之间以及之间的变异性和可重复性。使用标准化的双重SMAD抑制方法，所有iPSC系都产生了神经祖细胞，这些神经祖细胞随后可以分化为皮层神经元。通过免疫细胞化学定性分析神经分化，并通过qPCR定量分析祖细胞，神经元，皮层和神经胶质标志物。综上所述，我们提出了一个多步骤质量控制工作流程，以评估iPSC之间以及之间的变异性和可重复性。使用标准化的双重SMAD抑制方法，所有iPSC系都产生了神经祖细胞，这些神经祖细胞随后可以分化为皮层神经元。通过免疫细胞化学定性分析神经分化，并通过qPCR定量分析祖细胞，神经元，皮层和神经胶质标志物。综上所述，我们提出了一个多步骤质量控制工作流程，以评估iPSC之间以及之间的变异性和可重复性。