Guidelines for mouse and human DC generation.

Guidelines for mouse and human DC generation.
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DOI:
10.1002/eji.202249816
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发表时间:
2023-11
影响因子:
5.4
通讯作者:
--
中科院分区:
医学3区
文献类型:
--
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这篇文章是树突状细胞指南文章系列的一部分,该系列提供了一系列最先进的方案,用于制备,流式细胞术表型分析,生成,荧光显微镜,以及来自淋巴器官和各种非淋巴组织的小鼠和人类树突状细胞(DC)的功能表征。本文提供了从不同细胞来源(如鼠BM和HoxB 8细胞,以及来自脐带血、BM和外周血或外周血单核细胞的人CD 34+细胞)制备和成功生成小鼠和人DC的方案,其中包括最重要的细节和陷阱。我们描述了小鼠cDC 1,cDC 2,和pDC的生成与Flt 3L和BM衍生的DC与GM-CSF的生成。用于人DC生成的方案集中于在0 P9细胞层上的CD 34+细胞培养,用于cDC 1、cDC 2、cDC 3和pDC亚群生成以及来自外周血单核细胞(MoDC)的DC生成。另外的方案包括鼠DC亚群的富集、CRISPR/Cas9编辑和临床级人DC生成。虽然所有协议都是由经验丰富的科学家编写的,他们在工作中经常使用它们,但这篇文章也由领先的专家进行了同行评审,并得到了所有合著者的批准,使其成为基础和临床DC免疫学家的重要资源。
This article is part of the Dendritic Cell Guidelines article series, which provides a collection of state-of-the-art protocols for the preparation, phenotype analysis by flow cytometry, generation, fluorescence microscopy, and functional characterization of mouse and human dendritic cells (DC) from lymphoid organs and various non-lymphoid tissues. This article provides protocols with top ticks and pitfalls for preparation and successful generation of mouse and human DC from different cellular sources, such as murine BM and HoxB8 cells, as well as human CD34+ cells from cord blood, BM, and peripheral blood or peripheral blood monocytes. We describe murine cDC1, cDC2, and pDC generation with Flt3L and the generation of BM-derived DC with GM-CSF. Protocols for human DC generation focus on CD34+ cell culture on OP9 cell layers for cDC1, cDC2, cDC3, and pDC subset generation and DC generation from peripheral blood monocytes (MoDC). Additional protocols include enrichment of murine DC subsets, CRISPR/Cas9 editing, and clinical grade human DC generation. While all protocols were written by experienced scientists who routinely use them in their work, this article was also peer-reviewed by leading experts and approved by all co-authors, making it an essential resource for basic and clinical DC immunologists.
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