在巨细胞和B细胞中分析LC3相关的细胞和抗原呈现
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Svenja Luisa Nopper1, Katarina Wendy Schmidt1, Laure-Anne Ligeon1
1Viral Immunobiology, Institute of Experimental Immunology, University of Zürich, Zürich, Switzerland.
Methods in molecular biology (Clifton, N.J.)
|June 26, 2023
概括
与LC3相关的细胞分裂 (LAP) 和自对于对CD4+T细胞的抗原呈现至关重要. 这项研究详细介绍了研究B细胞和巨细胞中LAP和自的方法,以更好地了解免疫反应.
科学领域:
- 免疫学 免疫学 免疫学
- 细胞生物学 细胞生物学
- 分子生物学分子生物学
背景情况:
- 自和LC3-关联细胞分裂 (LAP) 对于通过MHCII类分子对CD4+T细胞进行抗原处理和呈现至关重要.
- 虽然它们在巨细胞和树突细胞中的作用已知,但它们在抗原处理期间在B细胞中的功能仍然不太了解.
研究的目的:
- 描述监测,操纵和理解LAP和古典自在人类单细胞衍生巨细胞和B细胞淋巴细胞状细胞系 (LCLs) 的抗原呈现中的作用的方法.
- 为产生人类细胞衍生原始巨细胞和LCLs提供协议.
主要方法:
- 使用CRISPR/Cas9进行ATG4B沉默的基因操纵和用于ATG4B过度表达的lentivirus传递.
- 触发LAP和量化ATG蛋白质的方法通过西方斑点和免疫光.
- 在体外共同培养试验通过测量CD4+T细胞分泌的细胞因子来评估MHCII类抗原呈现.
主要成果:
- 建立了产生人类单细胞衍生的巨细胞和LCLs的协议.
- 证明了操纵自途径 (ATG4B沉默/过度表达) 和触发LAP.的方法.
- 通过CD4+T细胞细胞因子生产来测量MHCII类抗原呈现的试验得到了验证.
结论:
- 描述的方法允许全面调查LAP和自细胞在巨细胞和B细胞的抗原呈现中.
- 这项工作为剖析这些途径对适应性免疫反应的贡献提供了必要的工具.
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