同位素标记相对和绝对定量技术研究进展

定量蛋白质组学是蛋白质研究的前沿学科。目前常用的定量蛋白质组学研究技术有荧光差异凝胶电泳（DIGE）、同位素亲和标记（ICAT）等。同位素标记相对和绝对定量（iTRAQ）技术是近年来最新开发的一种新的蛋白质组学定量研究技术。结合非凝胶串联质谱技术,该技术可对复杂样本、细胞器、细胞裂解液等样本进行相对和绝对定量研究,具有较好的定量效果、较高的重复性,并可对多达四种不同样本同时进行定量分析。本文对 iTRAQ 技术的原理、实验方法及应用进展进行了综述。     

高精度相对和绝对定量的等量异位标签在定量蛋白质组学研究中的新进展

蛋白质组学逐渐从定性研究转向定量研究。在定量蛋白质组学技术中,相对和绝对定量的等量异位标签(Isobaric tags for relative and absolute quantitation,iTRAQ)是应用最广泛的技术之一,具有通量高、稳定性强及不受样品来源制约等优点,几乎可以对任意样品进行标记,而且可以同时对多达8个样品进行定量分析,有效地提高了通量。iTRAQ技术不断改进,其定量准确性显著提高,适用的平台越来越多,为微生物、动物、植物、生物医学领域蛋白质及其翻译后修饰组研究创造了条件。文中综述了高精度iTRAQ技术在定量蛋白质组学研究中的最新发展及其应用。     

稳定同位素标记试剂DiART在蛋白质组中的定量特征研究

等重同位素标记方法,如同位素标记相对和绝对定量(iTRAQ),可以对肽段进行化学标记,之后通过质谱分析得到的报告离子的强度信息而实现对肽段的定量.目前,这种标记技术在定量蛋白质组学研究中有广泛应用.DiART也是一种等重同位素标记方法,且与iTRAQ的定量原理类似,但是其化学结构组成与iTRAQ有所不同,因而其定量特征有其独特表现.本研究通过对DiART标记的简单蛋白样品、复杂蛋白样品以及复杂样品中目标蛋白的定量情况进行了分析,从而对DiART的定量特征进行了研究,并同时与iTRAQ进行了比较.结果表明,DiART方法在定量稳定性、准确性及动态范围方面均更具优势.     

iTRAQ技术在医药领域的研究进展

蛋白质组学的研究是生命科学进入后基因时代的特征,是探索生命奥秘的有效手段,相关的实验技术已被广泛应用于各个领域。传统的蛋白组学技术包括凝胶电泳、质谱分析、蛋白质芯片等,但由于这些技术存在灵敏度低、操作繁琐、可重复性差等缺点已无法适应当今蛋白质研究的需要;这就促使了蛋白组学技术的改进和革新。同位素标记的相对和绝对定量(iTRAQ)技术是目前进行蛋白定性定量常用的、发展较快的新技术,该技术灵敏度高、准确率和可重复性好,能够同时检测多个样本,并且对样本进行精准高效的分析。文章梳理了iTRAQ技术的优缺点,对该技术近几年在医药基础和临床研究应用的情况进行整理和分析,探讨该技术在医药领域的应用前景。     

iTRAQ技术在真菌研究中的应用进展

iTRAQ技术是一种新的、功能强大的、可以最多同时比较8种不同样品中蛋白质相对或绝对含量的蛋白质组学方法,结合多维液相色谱和串联质谱分析,iTRAQ技术已成为差异蛋白质组学定量研究的主要工具之一。而真菌的致病作用是多种蛋白质共同参与的真菌?宿主相互作用的复杂过程,因此整体、定量地分析真菌致病过程中的差异表达蛋白质谱,对于研究真菌的致病机制具有重要作用。该文重点就iTRAQ技术在真菌研究中的应用进展进行综述。     

iTRAQ技术在动物生产中的应用

近年来,蛋白质组学的迅速发展使得相关实验技术也突飞猛进。与其他技术相比,同位素标记相对和绝对定量(isobaric tags for relative and absolute quantification,i TRAQ)技术以其高通量、高灵敏度、高重复性和能对各种复杂样品进行相对和绝对定量研究等优点迅速被广大学者接受,成为蛋白质组学定量研究的主要手段之一。i TRAQ技术的不断优化,使得它在动物、微生物等领域得到广泛应用。该文主要介绍了iTRAQ技术的基本原理和优缺点,并对i TRAQ技术在肉牛肌肉发育及品质中的作用进行了分析;介绍了奶牛乳蛋白质差异分析及乳房炎防御蛋白质筛选;介绍了绵羊产肉性能、毛囊发育及肠道免疫;介绍了生猪骨骼肌发育,猪圆环病毒II型(porcine circo virus type 2,PCV2)疾病防御,并进行冻存精液的影响因素的分析;介绍了肉鸡肌肉发育;同时也对蛋壳品质及鸡传染性法氏囊病(infectious bursal disease,IBD)防御机制研究等方面的研究进展进行了综述,以期为畜牧科学研究和生产实践提供依据。     

基于质谱的定量蛋白质组学策略和方法研究进展

定量蛋白质组学已经成为组学领域研究的热点之一.相关实验技术和计算方法的不断创新极大地促进了定量蛋白质组学的飞速发展.常用的定量蛋白质组学策略按照是否需要稳定同位素标记可以分为无标定量和有标定量两大类.每类策略又产生了众多定量方法和工具,它们一方面推动了定量蛋白质组学的深入发展;另一方面,也在实验策略与技术的发展过程中不断更新.因此对这些定量实验策略和方法进行系统总结和归纳将有助于定量蛋白质组学的研究.本文主要从方法学角度全面归纳了目前定量蛋白质组学研究的相关策略和算法,详述了无标定量和有标定量的具体算法流程并比较了各自特点,还对以研究蛋白质绝对丰度为目标的绝对定量算法进行了总结,列举了常用的定量软件和工具,最后概述了定量结果的质量控制方法,对定量蛋白质组学方法发展的前景进行了展望.     

植物蛋白质组学研究若干重要进展

植物蛋白质组学近年来正从定性向精确定量蛋白质组学的方向发展。国际上近两年发表的约160篇研究论文报道了利用不断改进的双向电泳结合生物质谱技术、多维蛋白质鉴定技术,以及包括双向荧光差异凝胶电泳、幅N体内代谢标记、同位素标记的亲和标签、同位素标记相对和绝对定量等在内的第2代蛋白质组学技术,对植物组织（器官）与细胞器、植物发育过程和植物响应环境胁迫的蛋白质组特征,以及植物蛋白质翻译后修饰和蛋白质相互作用等方面的研究成果。该文对上述报道进行总结,综述了2007年以来植物蛋白质组学若干重要问题研究的新进展。     

植物蛋白质组学研究若干重要进展

植物蛋白质组学近年来正从定性向精确定量蛋白质组学的方向发展。国际上近两年发表的约160篇研究论文报道了利用不断改进的双向电泳结合生物质谱技术、多维蛋白质鉴定技术, 以及包括双向荧光差异凝胶电泳、15N体内代谢标记、同位素标记的亲和标签、同位素标记相对和绝对定量等在内的第2代蛋白质组学技术, 对植物组织(器官)与细胞器、植物发育过程和植物响应环境胁迫的蛋白质组特征, 以及植物蛋白质翻译后修饰和蛋白质相互作用等方面的研究成果。该文对上述报道进行总结, 综述了2007年以来植物蛋白质组学若干重要问题研究的新进展。     

18O标记法在定量蛋白质组学中的应用

基于质谱技术去识别和检测蛋白质表达差异是一个热点,有助于生物过程和体系的分子机制的研究。近年来各种基于质谱技术的定量蛋白质组学研究方法发展较快,相对其他方法而言,18O标记法是一种较为理想、相对容易实现并且在不断完善的体外标记方法,最近在定量蛋白质组学研究中应用较多。现对18O标记法原理、特点以及技术方法的优化和应用进展进行综述。     

Proteomics in cancer vaccine development

Proteomics is a new scientific field aimed at the large-scale characterization of the protein constituents of biologic systems. It facilitates comparisons between different protein preparations by searching for minute differences in their protein expression repertoires and the patterns of their post-translational modifications. These attributes make proteomics perfectly suited for searching for proteins and peptides expressed exclusively or preferentially in cancer cells as candidates for cancer vaccines. The main proteomics technologies include 2D polyacrylamide gel electrophoresis, multidimensional high-performance liquid chromatography, mass spectrometry and protein arrays. Proteomics technologies used to analyze cancer culture cells, fresh tumor specimens, human leukocyte antigen peptides, serum and serum antibodies (serologic proteomics) have successfully identified tumor markers. Turning the potential vaccine candidates identified by proteomics technologies into clinical treatments awaits demonstration.     

Driving biochemical discovery with quantitative proteomics

Proteomic analysis of biological samples plays an increasing role in modern research. Although the application of proteomics technologies varies across many disciplines, proteomics largely is a tool for discovery that then leads to novel hypotheses. In recent years, new methods and technologies have been developed and applied in many areas of proteomics, and there is a strong push towards using proteomics in a quantitative manner. Indeed, mass spectrometry-based, quantitative proteomics approaches have been applied to great success in a variety of biochemical studies. In particular, the use of quantitative proteomics provides new insights into protein complexes and post-translational modifications and leads to the generation of novel insights into these important biochemical systems.     

基于质谱的植物蛋白质组学研究方法

基于质谱的植物蛋白质组学研究方法,从定性和定量蛋白质组学两个方向进行了归纳总结,并对近年来出现的靶向蛋白质组学、DIA/SWATH技术、化学蛋白质组学,以及多组学联合分析等蛋白质组学研究的新技术、新方法和新应用进行了综述。     

An integrated proteomic workflow for two-dimensional differential gel electrophoresis and robotic spot picking

New technologies have advanced the field of proteomics, and a number of companies have developed innovative platforms to drive this research. However, significant challenges are often encountered when trying to integrate complementary technologies from multiple manufacturers. We have developed a software and hardware solution to integrate the Ettan two-dimensional difference gel electrophoresis (2-D DIGE) system (GE Healthcare) with the Investigator ProPic spot picking robot (Genomic Solutions). We have analyzed protein sample preparations from bacterial and mammalian sources to demonstrate a new workflow with increased throughput for gel-based proteomics.     

Proteomics technologies and challenges

Proteomics is the study of proteins and their interactions in a cell. With the completion of the Human Genome Project, the emphasis is shifting to the protein compliment of the human organism. Because proteome reflects more accurately on the dynamic state of a cell, tissue, or organism, much is expected from proteomics to yield better disease markers for diagnosis and therapy monitoring. The advent of proteomics technologies for global detection and quantitation of proteins creates new opportunities and challenges for those seeking to gain greater understanding of diseases. High-throughput proteomics technologies combining with advanced bioinformatics are extensively used to identify molecular signatures of diseases based on protein pathways and signaling cascades. Mass spectrometry plays a vital role in proteomics and has become an indispensable tool for molecular and cellular biology. While the potential is great, many challenges and issues remain to be solved, such as mining low abundant proteins and integration of proteomics with genomics and metabolomics data. Nevertheless, proteomics is the foundation for constructing and extracting useful knowledge to biomedical research. In this review, a snapshot of contemporary issues in proteomics technologies is discussed.     

Oncoproteomics: current trends and future perspectives

Oncoproteomics is the application of proteomics technologies in oncology. Functional proteomics is a promising technique for the rational identification of biomarkers and novel therapeutic targets for cancers. Recent progress in proteomics has opened new avenues for tumor-associated biomarker discovery. With the advent of new and improved proteomics technologies, such as the development of quantitative proteomic methods, high-resolution, -speed and -sensitivity mass spectrometry and protein arrays, as well as advanced bioinformatics for data handling and interpretation, it is now possible to discover biomarkers that can reliably and accurately predict outcomes during cancer management and treatment. However, there are several difficulties in the study of proteins/peptides that are not inherent in the study of nucleic acids. New challenges arise in large-scale proteomic profiling when dealing with complex biological mixtures. Nevertheless, oncoproteomics offers great promise for unveiling the complex molecular events of tumorigenesis, as well as those that control clinically important tumor behaviors, such as metastasis, invasion and resistance to therapy. In this review, the development and advancement of oncoproteomics technologies for cancer research in recent years are expounded.     

Oncoproteomics: current trends and future perspectives

Oncoproteomics is the application of proteomics technologies in oncology. Functional proteomics is a promising technique for the rational identification of biomarkers and novel therapeutic targets for cancers. Recent progress in proteomics has opened new avenues for tumor-associated biomarker discovery. With the advent of new and improved proteomics technologies, such as the development of quantitative proteomic methods, high-resolution, -speed and -sensitivity mass spectrometry and protein arrays, as well as advanced bioinformatics for data handling and interpretation, it is now possible to discover biomarkers that can reliably and accurately predict outcomes during cancer management and treatment. However, there are several difficulties in the study of proteins/peptides that are not inherent in the study of nucleic acids. New challenges arise in large-scale proteomic profiling when dealing with complex biological mixtures. Nevertheless, oncoproteomics offers great promise for unveiling the complex molecular events of tumorigenesis, as well as those that control clinically important tumor behaviors, such as metastasis, invasion and resistance to therapy. In this review, the development and advancement of oncoproteomics technologies for cancer research in recent years are expounded.     

Nutriproteomics: identifying the molecular targets of nutritive and non-nutritive components of the diet

The study of whole patterns of changes in protein expression and their modifications, or proteomics, presents both technological advances as well as formidable challenges to biological researchers. Nutrition research and the food sciences in general will be strongly influenced by the new knowledge generated by the proteomics approach. This review examines the different aspects of proteomics technologies, while emphasizing the value of consideration of "traditional" aspects of protein separation. These include the choice of the cell, the subcellular fraction, and the isolation and purification of the relevant protein fraction (if known) by protein chromatographic procedures. Qualitative and quantitative analyses of proteins and their peptides formed by proteolytic hydrolysis have been substantially enhanced by the development of mass spectrometry technologies in combination with nanoscale fluidics analysis. These are described, as are the pros and cons of each method in current use.     

A Bioinformatics Perspective on Proteomics: Data Storage, Analysis, and Integration

The field of proteomics is advancing rapidly as a result of powerful new technologies and proteomics experiments yield a vast and increasing amount of information. Data regarding protein occurrence, abundance, identity, sequence, structure, properties, and interactions need to be stored. Currently, a common standard has not yet been established and open access to results is needed for further development of robust analysis algorithms. Databases for proteomics will evolve from pure storage into knowledge resources, providing a repository for information (meta-data) which is mainly not stored in simple flat files. This review will shed light on recent steps towards the generation of a common standard in proteomics data storage and integration, but is not meant to be a comprehensive overview of all available databases and tools in the proteomics community.     

A proteomics strategy for the analysis of bacterial biodegradation pathways

Bacterial biodegradation (bioremediation) is the use of microorganisms to break down organic materials into simpler compounds; it plays a pivotal role in the clean-up of hazardous wastes in the environment. Following the completion of genome sequencing in bacteria capable of biodegradation, functional genomic studies have played a major role in obtaining information on bacterial biodegradation pathways. Novel proteomics technologies have recently been developed to make it possible to analyze global protein expression. Proteomics can also provide important information on the life cycle, regulation, and post-translational modification of proteins induced under specific conditions. Proteomics technologies have been applied to the comprehensive study of bacterial biodegradation. In this paper, we introduce the proteomics technologies applicable to bacterial biodegradation studies, review the results of the proteomics analysis of representative biodegrading bacteria, and discuss the potential use of proteomics technologies in future biodegradation studies.