使用CRISPR-Cas9技术在THP-1细胞中研究内源性产生的伊塔科纳酸的作用的协议
Luke A Bourner1, Katie A Acken1, Haiyan Long1
1Eli Lilly and Company, Lilly Corporate Center, Indianapolis, IN, USA.
STAR protocols
|January 23, 2026
概括
研究人员开发了一种CRISPR-Cas9方法,以消除免疫细胞中产生伊塔康酸盐的酶 (ACOD1). 这种技术可以更好地研究内源性伊塔康酸盐,这是炎症的关键调节剂.
科学领域:
- 免疫学 免疫学 免疫学
- 分子生物学分子生物学
- 生物化学 生物化学
背景情况:
- 内源性伊塔科纳酸调节先天性免疫反应和促炎性细胞因子.
- 现有的伊塔科纳特模仿剂不能完全复制内源性伊塔科纳特的功能.
研究的目的:
- 开发一种CRISPR-Cas9协议,用于在THP-1细胞中切除产生伊塔康酸盐的酶ACOD1.
- 为了研究内源性伊塔科纳酸在免疫调节中的作用.
主要方法:
- 在CRISPR-Cas9基因编辑中删除IRG1基因中的特定区域.
- 免疫洗,ELISA和液体染色学-质谱学 (LC-MS) 用于功能验证.
- 在修改的THP-1细胞中量化伊塔科纳酸盐水平.
主要成果:
- 在IRG1中成功删除了一个4核酸区,从而消除了ACOD1的功能.
- 通过多种生物化学和免疫学测定验证了ACOD1删除.
- 建立了一种研究THP-1巨细胞内源性伊塔康酸盐的方法.
结论:
- 开发的CRISPR-Cas9协议有效地消除了ACOD1,从而可以精确地研究内源性伊塔科纳酸.
- 这种方法为研究伊塔科纳酸在先天免疫和炎症中的作用提供了有价值的工具.
- 促进针对伊塔康酸路径的免疫调节疗法的研究.
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