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{"title":"Purification of Live Stem-Cell-Derived Islet Lineage Intermediates","authors":"Niloofar Rasouli, Douglas A. Melton, Juan R. Alvarez-Dominguez","doi":"10.1002/cpsc.111","DOIUrl":null,"url":null,"abstract":"<p>Stem-cell-derived tissues offer platforms to study organ development, model physiology during health and disease, and test novel therapies. We describe methods to isolate cells at successive stages during in vitro differentiation of human stem cells into the pancreatic endocrine lineage. Using flow cytometry, we purify live lineage intermediates in numbers not available by fetal biopsy. These include pancreatic and endocrine progenitors, isolated based on known surface markers. We further report a strategy that leverages intracellular zinc content and DPP4/CD26 expression to separate monohormonal insulin<sup>+</sup> β cells from polyhormonal counterparts. These methods enable comprehensive molecular profiling during human islet lineage progression. © 2020 Wiley Periodicals LLC.</p><p><b>Basic Protocol</b>: In vitro isolation of human islet developmental intermediates</p>","PeriodicalId":53703,"journal":{"name":"Current Protocols in Stem Cell Biology","volume":"53 1","pages":""},"PeriodicalIF":0.0000,"publicationDate":"2020-06-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpsc.111","citationCount":"3","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Current Protocols in Stem Cell Biology","FirstCategoryId":"1085","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1002/cpsc.111","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"Biochemistry, Genetics and Molecular Biology","Score":null,"Total":0}
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Abstract
Stem-cell-derived tissues offer platforms to study organ development, model physiology during health and disease, and test novel therapies. We describe methods to isolate cells at successive stages during in vitro differentiation of human stem cells into the pancreatic endocrine lineage. Using flow cytometry, we purify live lineage intermediates in numbers not available by fetal biopsy. These include pancreatic and endocrine progenitors, isolated based on known surface markers. We further report a strategy that leverages intracellular zinc content and DPP4/CD26 expression to separate monohormonal insulin+ β cells from polyhormonal counterparts. These methods enable comprehensive molecular profiling during human islet lineage progression. © 2020 Wiley Periodicals LLC.
Basic Protocol : In vitro isolation of human islet developmental intermediates
活干细胞衍生胰岛谱系中间体的纯化
干细胞衍生的组织为研究器官发育、健康和疾病期间的生理模型以及测试新疗法提供了平台。我们描述了在人类干细胞体外分化成胰腺内分泌谱系的连续阶段分离细胞的方法。使用流式细胞术，我们纯化了活体细胞系中间体，其数量无法通过胎儿活检获得。这些包括胰腺和内分泌祖细胞，根据已知的表面标记分离。我们进一步报道了一种利用细胞内锌含量和DPP4/CD26表达将单激素胰岛素+ β细胞与多激素胰岛素+ β细胞分离的策略。这些方法能够在人类胰岛谱系进展过程中进行全面的分子分析。©2020 Wiley期刊公司基本方案:体外分离人类胰岛发育中间体
本文章由计算机程序翻译，如有差异，请以英文原文为准。
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