{"title":"成体干细胞人肠道类器官的建立与培养","authors":"Cayetano Pleguezuelos-Manzano, Jens Puschhof, Stieneke van den Brink, Veerle Geurts, Joep Beumer, Hans Clevers","doi":"10.1002/cpim.106","DOIUrl":null,"url":null,"abstract":"<p>Human intestinal organoids derived from adult stem cells are miniature ex vivo versions of the human intestinal epithelium. Intestinal organoids are useful tools for the study of intestinal physiology as well as many disease conditions. These organoids present numerous advantages compared to immortalized cell lines, but working with them requires dedicated techniques. The protocols described in this article provide a basic guide to establishment and maintenance of human intestinal organoids derived from small intestine and colon biopsies. Additionally, this article provides an overview of several downstream applications of human intestinal organoids. © 2020 The Authors.</p><p><b>Basic Protocol 1</b>: Establishment of human small intestine and colon organoid cultures from fresh biopsies</p><p><b>Basic Protocol 2</b>: Mechanical splitting, passage, and expansion of human intestinal organoids</p><p><b>Alternate Protocol</b>: Differentiation of human intestinal organoids</p><p><b>Basic Protocol 3</b>: Cryopreservation and thawing of human intestinal organoids</p><p><b>Basic Protocol 4</b>: Immunofluorescence staining of human intestinal organoids</p><p><b>Basic Protocol 5</b>: Generation of single-cell clonal intestinal organoid cultures</p><p><b>Support Protocol 1</b>: Production of Wnt3A conditioned medium</p><p><b>Support Protocol 2</b>: Production of Rspo1 conditioned medium</p><p><b>Support Protocol 3</b>: Extraction of RNA from intestinal organoid cultures</p>","PeriodicalId":10733,"journal":{"name":"Current Protocols in Immunology","volume":"130 1","pages":""},"PeriodicalIF":0.0000,"publicationDate":"2020-09-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpim.106","citationCount":"60","resultStr":"{\"title\":\"Establishment and Culture of Human Intestinal Organoids Derived from Adult Stem Cells\",\"authors\":\"Cayetano Pleguezuelos-Manzano, Jens Puschhof, Stieneke van den Brink, Veerle Geurts, Joep Beumer, Hans Clevers\",\"doi\":\"10.1002/cpim.106\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p>Human intestinal organoids derived from adult stem cells are miniature ex vivo versions of the human intestinal epithelium. Intestinal organoids are useful tools for the study of intestinal physiology as well as many disease conditions. These organoids present numerous advantages compared to immortalized cell lines, but working with them requires dedicated techniques. The protocols described in this article provide a basic guide to establishment and maintenance of human intestinal organoids derived from small intestine and colon biopsies. Additionally, this article provides an overview of several downstream applications of human intestinal organoids. © 2020 The Authors.</p><p><b>Basic Protocol 1</b>: Establishment of human small intestine and colon organoid cultures from fresh biopsies</p><p><b>Basic Protocol 2</b>: Mechanical splitting, passage, and expansion of human intestinal organoids</p><p><b>Alternate Protocol</b>: Differentiation of human intestinal organoids</p><p><b>Basic Protocol 3</b>: Cryopreservation and thawing of human intestinal organoids</p><p><b>Basic Protocol 4</b>: Immunofluorescence staining of human intestinal organoids</p><p><b>Basic Protocol 5</b>: Generation of single-cell clonal intestinal organoid cultures</p><p><b>Support Protocol 1</b>: Production of Wnt3A conditioned medium</p><p><b>Support Protocol 2</b>: Production of Rspo1 conditioned medium</p><p><b>Support Protocol 3</b>: Extraction of RNA from intestinal organoid cultures</p>\",\"PeriodicalId\":10733,\"journal\":{\"name\":\"Current Protocols in Immunology\",\"volume\":\"130 1\",\"pages\":\"\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2020-09-17\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://sci-hub-pdf.com/10.1002/cpim.106\",\"citationCount\":\"60\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Current Protocols in Immunology\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://onlinelibrary.wiley.com/doi/10.1002/cpim.106\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"Immunology and Microbiology\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Current Protocols in Immunology","FirstCategoryId":"1085","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1002/cpim.106","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"Immunology and Microbiology","Score":null,"Total":0}
引用次数: 60
Establishment and Culture of Human Intestinal Organoids Derived from Adult Stem Cells
Human intestinal organoids derived from adult stem cells are miniature ex vivo versions of the human intestinal epithelium. Intestinal organoids are useful tools for the study of intestinal physiology as well as many disease conditions. These organoids present numerous advantages compared to immortalized cell lines, but working with them requires dedicated techniques. The protocols described in this article provide a basic guide to establishment and maintenance of human intestinal organoids derived from small intestine and colon biopsies. Additionally, this article provides an overview of several downstream applications of human intestinal organoids. © 2020 The Authors.
Basic Protocol 1: Establishment of human small intestine and colon organoid cultures from fresh biopsies
Basic Protocol 2: Mechanical splitting, passage, and expansion of human intestinal organoids
Alternate Protocol: Differentiation of human intestinal organoids
Basic Protocol 3: Cryopreservation and thawing of human intestinal organoids
Basic Protocol 4: Immunofluorescence staining of human intestinal organoids
Basic Protocol 5: Generation of single-cell clonal intestinal organoid cultures
Support Protocol 1: Production of Wnt3A conditioned medium
Support Protocol 2: Production of Rspo1 conditioned medium
Support Protocol 3: Extraction of RNA from intestinal organoid cultures
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