基于单颗粒计数的蛋白质定量方法研究进展

张馨艺; 米薇; 胡志上

doi:10.12338/j.issn.2096-9015.2023.0093

基于单颗粒计数的蛋白质定量方法研究进展

doi: 10.12338/j.issn.2096-9015.2023.0093

中国计量科学研究院，北京 100029

基金项目: 国家重点研发计划项目(2021YFF0600702)；中国计量科学研究院重点领域项目(AKYZD2114-1)。

详细信息

作者简介:
张馨艺（1998-），中国计量科学研究院在读研究生，研究方向：抗体药物分析技术与测量装置研究，邮箱：karmic_xy@163.com

通讯作者:
胡志上（1979-），中国计量科学研究院副研究员，研究方向：蛋白质化学计量，邮箱：huz@nim.ac.cn

中图分类号: TB98
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出版历程
- 收稿日期: 2023-03-29
- 录用日期: 2023-04-03
- 修回日期: 2023-05-21
- 网络出版日期: 2023-06-29
- 刊出日期: 2023-04-18

Research Progress in Protein Quantification Based on Single Particle Counting in Large-Scale Metrology

National Institute of Metrology, Beijing 100029, China

摘要

摘要: 蛋白质是执行功能的重要的生物大分子，蛋白质类生物标志物的鉴别与定量和复杂基质中目标蛋白准确含量的测定对疾病的预防、诊断及治疗有重要意义。伴随着生命科学的不断发展，科学家开发出许多种蛋白质定量方法，较为常见的有紫外可见分光光度法、酶联免疫吸附法及基于质谱的定量方法。但是对于高分子量蛋白质，尤其是分子量在200 kDa以上的蛋白质，由于其结构复杂及分子量巨大，导致前处理复杂或反应不专一、不完全，造成定量结果不准确。单颗粒计数法是通过对溶液中粒子逐一计数实现定量，相比于传统定量方法，单颗粒计数法具有灵敏度高、区分度好、外界因素干扰少的特点，加之此类方法样品处理简单，属于非破坏性的分析，适合对高分子量蛋白质进行完整结构水平的定量。总结了蛋白质定量方法，重点关注基于单颗粒计数的3种典型技术，包括单颗粒电感耦合等离子体质谱法（SP-ICP-MS）、单分子阵列技术（Simoa）和电喷雾-差分电迁移率-凝结核颗粒计数法（ES-DMA-CPC），系统阐述了这几种方法的工作原理及在蛋白质定量中的应用，并对未来该领域的发展进行了展望。
- 计量学 /
- 高分子量蛋白质 /
- 单颗粒计数 /
- 定量分析 /
- 单颗粒电感耦合等离子体质谱法 /
- 单分子阵列技术 /
- 电喷雾-差分电迁移率-凝结核颗粒计数法
Abstract: Proteins, as crucial biological macromolecules, perform essential functions. The identification and quantification of protein biomarkers, as well as accurate measurement of target proteins in complex matrices, are vital for disease prevention, diagnosis, and treatment. With the ongoing development of life sciences, scientists have introduced several protein quantification methods, including ultraviolet-visible spectrophotometry, enzyme-linked immunosorbent assay, and mass spectrometry-based quantitative methods. However, for high molecular weight proteins, especially those exceeding 200 kDa, their complex structure and enormous molecular weight lead to complicated preprocessing or non-specific, incomplete reactions, causing inaccurate quantification results. Single particle counting quantifies by individually counting particles in a solution. This method offers higher sensitivity, better differentiation, and less interference than traditional methods. Furthermore, given its simple sample preparation and non-destructive nature of analysis, single particle counting is well-suited for quantifying high molecular weight proteins at the complete structural level. This paper provides a comprehensive summary of protein quantification methods, with a specific focus on three typical single particle counting techniques: single particle inductively coupled plasma mass spectrometry (SP-ICP-MS), single molecule array (Simoa), and electrospray differential mobility analysis-condensation particle counter (ES-DMA-CPC). The operating principles of these methods and their applications in protein quantification are systematically explained, and the paper concludes with a projection of future developments in the field of protein quantification based on single particle counting.
- metrology /
- high molecular weight protein /
- single particle counting /
- quantitative analysis /
- SP-ICP-MS /
- Simoa /
- ES-DMA-CPC

HTML全文

图 1 SP-ICP-MS工作示意图

Figure 1. Schematic of SP-ICP-MS operation

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图 2 Simoa工作示意图

Figure 2. Schematic of Simoa operation

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图 3 ES-DMA-CPC工作示意图

Figure 3. Schematic of ES-DMA-CPC operation

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表 1 本文中提到的基于单颗粒计数的蛋白质定量方法及优缺点概述

Table 1. Overview of the advantages and disadvantages of protein quantification methods based on single particle counting discussed in this paper

	优点	缺点
SP-ICP-MS	多种标签可用于定量，特异性高，信噪比高，分析速度快	不同操作者标记效率差异较大，传输效率无法准确测定，杂质峰干扰
Simoa	灵敏度极高，检出限低，分析速度快，线性范围宽	交叉反应，非特异性结合，背景干扰
ES-DMA-CPC	无需标记，不破坏样品，准确度高	随机误差较大，误差来源研究不充分

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