蛋白酶激活受体的特征调节器与调动试验使用盘子阅读器
Jacob T DeRousse1, Chris Dockendorff2
1Function Therapeutics; Department of Chemistry, Marquette University; School of Medicine, Washington University.
Journal of visualized experiments : JoVE
|June 10, 2024
概括
这项研究提出了一种具有成本效益的板读器测试,用于测量细胞变化,从而能够对蛋白酶激活受体 (PAR) 连接体和Gαq信号激活进行中速选.
科学领域:
- 细胞生物学 细胞生物学
- 生物化学 生化学
- 药理学 药理学是指药理学的学科.
背景情况:
- 高通量细胞监测使用昂贵的专用成像仪器.
- 广泛可用的板读器提供了一个更容易获得的替代方案,但需要测试优化.
- 内皮细胞系是研究信号通路的关键模型.
研究的目的:
- 开发和优化用于测量动员的中等吞吐量板读取器试验.
- 评估蛋白酶激活受体 (PAR) 信号传递和Gαq通路激活.
- 为了能够选PAR配体,包括新型抗炎化合物.
主要方法:
- 使用了一个内皮细胞系 (EA.hy926) 在一个96井板格式.
- 使用结合染料Fluo-4来监测细胞内度变化.
- 通过PAR受体激活触发的测量Gαq信号和动员.
- 优化测试条件,避免需要自动化液体处理.
主要成果:
- 成功建立了一个优化的盘子阅读器测试,用于量化动员.
- 证明了该试验能够测量 PAR 驱动的 Gαq 信号激活.
- 描述了各种PAR配体,包括全性PAR1向的抗炎性配体.
- 在不需要自动化液体处理器的情况下验证了测试的适用性.
结论:
- 开发的板阅读器测试提供了一种具有成本效益和可访问的方法,用于PAR配体的中等通量选.
- 该协议有助于研究内皮细胞中Gαq信号传递和调动.
- 该测试可用于研究诱导信号通路的其他受体.
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