Optimized, Efficient Measurement of the Expression of Undifferentiated Stem Cell Markers in Human Induced Pluripotent Stem Cells (iPSCs) by Flow Cytometry

Current protocols Pub Date : 2025-03-03 DOI:10.1002/cpz1.70105

Vaishanavi Saware, Wendy Runyon, Sam Hu, Benjamin van Soldt, Ritu Kumar, Jane Srivastava

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Abstract

Induced pluripotent stem cells (iPSCs) have revolutionized the fields of regenerative medicine, disease modeling, and drug discovery. However, the usage of iPSCs for various applications has been hampered by the observed line-to-line variability in their differentiation capacity. Therefore, it is important to verify the pluripotent status of iPSCs. A very effective way to define the pluripotent state of iPSCs is by evaluating the expression of established undifferentiated stem cell markers. A bona fide iPSC must have high, homogeneous expression of these markers. Here, we present a cost-effective platform that can be readily utilized by researchers to define the pluripotency status of iPSCs by measuring the expression of surface and intracellular markers by flow cytometry. © 2025 Wiley Periodicals LLC.

Basic Protocol 1: iPSC culture and collection for flow cytometry analysis

Basic Protocol 2: Staining of iPSCs for extracellular and intracellular undifferentiated stem cell markers

Basic Protocol 3: Flow cytometry acquisition

Basic Protocol 4: Flow cytometry data analysis

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诱导多能干细胞（iPSC）为再生医学、疾病建模和药物发现领域带来了革命性的变化。然而，由于观察到 iPSCs 的分化能力存在品系间差异，其在各种应用中的使用受到了阻碍。因此，验证 iPSCs 的多能状态非常重要。确定 iPSCs 多能状态的一个非常有效的方法是评估已建立的未分化干细胞标记物的表达。真正的 iPSC 必须具有这些标志物的高均匀表达。在这里，我们提出了一个经济高效的平台，研究人员可随时利用该平台，通过流式细胞术测量细胞表面和细胞内标记物的表达，从而确定 iPSC 的多能状态。© 2025 Wiley Periodicals LLC.基本方案1：iPSC培养和收集用于流式细胞仪分析基本方案2：iPSC细胞外和细胞内未分化干细胞标记物染色基本方案3：流式细胞仪采集基本方案4：流式细胞仪数据分析

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Current protocols

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4.00

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