开发一种自动化,高通量方法,用于原生质谱和碰撞诱导的展开
Brock R Juliano1, Joseph W Keating1, Henry W Li1
1Department of Chemistry, University of Michigan, Ann Arbor, Michigan 48109, United States.
Analytical chemistry
|November 4, 2023
概括
我们开发了一种使用RapidFire机器人样本处理的自动化高通量方法,用于本地离子移动性质谱 (nIM-MS) 和碰撞诱导展开 (CIU). 这种方法显著增加了生物分子结构和稳定性分析的吞吐量.
科学领域:
- 生物物理化学 生物物理化学
- 分析化学 分析化学
- 结构生物学 结构生物学
背景情况:
- 原生离子移动性质谱 (nIM-MS) 对于快速评估生物分子结构非常有价值.
- 碰撞诱导展开 (CIU) 实验通过提供对生物分子构造和稳定性的洞察来增强nIM-MS.
- 目前的nIM-MS和CIU方法受到缓慢,非自动化样本准备和引入的限制.
研究的目的:
- 使用机器人样本处理系统开发用于本地质谱 (nMS) 和CIU的自动化,高通量方法.
- 建立本地RapidFire-MS (nRapidFire-MS) 以快速在线淡化和样品引入.
- 为了证明nRapidFire-MS对于高通量CIU分析的实用性.
主要方法:
- 使用RapidFire机器人样品处理系统进行自动样品准备和引入.
- 通过使用尺寸排除色谱弹实施在线淡盐处理.
- 采集了使用nRapidFire-MS系统的碰撞诱导展开 (CIU) 指纹.
主要成果:
- 通过nRapidFire-MS. 在每样仅需10秒的时间内实现样品引入.
- 收集的CIU指纹在30秒内脱盐后.
- 与标准的nMS和CIU相比,nRapidFire-MS数据显示了相同的离子碰撞截面,证实了精确的构造性人口跟踪.
- 已证明适用于各种生物分子类,包括蛋白质 (5300 kDa),蛋白质复合体和寡核酸.
- nRapidFire-MS显示了生物制药样品高通量分析的潜力.
结论:
- 开发的nRapidFire-MS方法使得高通量本地质谱和CIU分析成为可能.
- 这种自动化方法克服了传统方法的吞吐量限制.
- nRapidFire-MS具有促进蛋白质工程,抑制剂发现和生物治疗配方开发的巨大潜力.
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