高性能本地分离mAb蛋白质形式的CZE-MS在本地和变质纳米ESI条件下超敏感电荷变体表征
Ann-Katrin Schwenzer1, Christian Neusüß1
1Faculty of Chemistry, Aalen University, Beethovenstraße 1, Aalen 73430, Germany.
Analytical chemistry
|August 11, 2025
概括
一种新的毛细管区电泳质谱法 (CZE-MS) 方法使治疗单克隆抗体 (mAb) 电荷变异的详细表征成为可能. 这种技术可以对蛋白质形式进行敏感的识别,这对于生物制药质量控制至关重要.
科学领域:
- 生物制药学分析
- 分析化学 分析化学
- 蛋白质组学是指蛋白质组学.
背景情况:
- 监测治疗抗体中的电荷变异对于确保药物的有效性和安全性至关重要.
- 毛细管区电泳 (CZE) 有效地分离电荷变体,但与质谱法 (MS) 不兼容,阻碍了变体识别.
- 现有的CZE方法用于电荷变量分析通常使用电解质,防止直接合到MS.
研究的目的:
- 开发和介绍一种新的CZE-MS方法,用于详细描述单克隆抗体 (mAb) 电荷变体和其他蛋白质形式.
- 为了实现与现有的非MS兼容方法相比较的分离性能,同时能够直接识别MS.
- 以高灵敏度和灵活性在完整水平上评估治疗mAbs的异质性.
主要方法:
- 在生理学pH下,使用中性静态毛细血管涂层与基于酸的电解质一起使用.
- 该CZE系统使用纳米流液体 (SL) 接口为MS进行了连接,用于超敏感的电离.
- 不同的SL在化和非化纳米ESI条件下进行了测试,以优化电离和光谱质量.
主要成果:
- 开发的CZE-MS方法证明了mAb电荷变体的高分离性能,类似于已建立的非MS兼容技术.
- 该方法使得大量蛋白质形式的敏感检测和鉴定成为可能:NISTmAb为115个,trastuzumab为70个.
- 确定了几种以前在完整水平上未被检测到的蛋白质形式,为mAb异质性提供了更深入的见解.
结论:
- 提出的CZE-MS方法提供了一个快速而详细的方法,用于在完整水平上评估治疗mAb异质性.
- 这种技术显著提高了生物制药的蛋白形溶解结构和功能表征的能力.
- 该方法克服了传统CZE的局限性,因为它允许MS直接识别分开的收费变体.
相关概念视频
Peptide Identification Using Tandem Mass Spectrometry
6.8K
Tandem mass spectrometry, also known as MS/MS or MS2, is an analytical technique that employs two mass analyzers. Essentially it is a series of mass spectrometers that helps isolate a particular biomolecule and then helps study its chemical properties.
This technique helps gather information regarding the protein from which the peptide was obtained and to study the peptides’ amino acid sequence. Identifying peptides from a complex mixture is an important component of the growing field of...
This technique helps gather information regarding the protein from which the peptide was obtained and to study the peptides’ amino acid sequence. Identifying peptides from a complex mixture is an important component of the growing field of...
6.8K
High-Resolution Mass Spectrometry (HRMS)
1.6K
The resolution of a mass spectrometer depends on the efficiency of separating ions with different ion masses. The mass of an atom is approximated to the sum of the masses of protons and neutrons inside, considering the masses of protons and neutrons as equal. However, the masses of the proton (1.6726 × 10−24 g) and neutron (1.6749 × 10−24 g) are not truly equal. There is a minor error in the expression of atomic masses relative to the simplest atom of hydrogen. For...
1.6K
Capillary Electrophoresis: Applications
530
Capillary electrophoretic separations offer various modes, each with unique applications. These modes include capillary zone electrophoresis, capillary gel electrophoresis, capillary array electrophoresis, capillary isoelectric focusing, capillary isotachophoresis, micellar electrokinetic chromatography, and capillary electrochromatography.
Capillary zone electrophoresis (CZE) separates ionic components based on their electrophoretic mobility. It has been used to separate proteins, amino acids,...
Capillary zone electrophoresis (CZE) separates ionic components based on their electrophoretic mobility. It has been used to separate proteins, amino acids,...
530
Mass Spectrometry: Complex Analysis
890
Mass spectrometry is an important technique for the identification of pure compounds. However, it has some limitations for the analysis of complex mixtures, often due to excessive fragmentation making the spectrum too complicated to decipher. Mass spectrometry can be combined with suitable separation methods in sequence, forming hyphenated methods, which are useful in the analysis of complex mixtures.
GC–MS is a powerful hyphenated method commonly used in forensics and environmental...
GC–MS is a powerful hyphenated method commonly used in forensics and environmental...
890
Mass Spectrometry: Overview
5.9K
Mass spectrometry is an analytical technique used to determine the molecular mass and molecular formula of a compound. The basic principle of mass spectrometry is to generate ions from the analyte molecule and measure these ion abundances against their molecular mass. One common type of ionization, known as electrospray ionization or EI, bombards the analyte molecules in the gas phase with high-energy electron beams. The electron beams displace an electron from the molecule and leave...
5.9K
Mass Analyzers: Overview
807
The mass analyzer is a crucial component of the mass spectrometer. In the ionization chamber, the vaporized sample is bombarded with a high-energy electron beam to generate a radical cation and further fragment into neutral molecules, radicals, and cations. A series of negatively charged accelerator plates accelerate the cations into the mass analyzer. The mass analyzer separates ions according to their mass-to-charge (m/z) ratios and then directs them to the detector. The common types of mass...
807


