Identity^E 高清蛋白质组系统

Identity^E High Definition Proteomics系统定义了严密的蛋白质识别的新标准体系。专门设计用于权威和全面的定性分析，沃特世解决方案使得蛋白质组学研究人员可参照色谱滞留时间、前体和产物离子块、结构意义、生物环境以及对多种肽段的识别，自信地识别蛋白质。

该Identity^E系统专门设计用于定性分析复杂的蛋白质消化，利用UPLC^®分离的高分辨率和新型的MS^E方法，提供卓越的色谱分析结果以及同步的前体和产物离子的信息。

该系统包括综合肽离子计数信息。该软件能够利用多层次的蛋白质初级结构的物化模型来帮助您想象和识别多肽和蛋白质，生成最高品质的MRM转换信息，用于串联四级生物标志物的验证和确认。

Identity^E系统能够提供基于情境的完整信息，用于自信地识别蛋白质—实现更有价值的生物学和医学研究。该系统适合那些正在寻找方法以改善定性蛋白质分析结果的可重复性和可靠性等的实验室，例如：

• 大学的生物/生化院系
• 核心蛋白质组学实验室
• 癌症研究设施
• 蛋白质的功能/交互作用实验室
• 生物技术公司

Identity^E高清蛋白质组学系统配置如下：

• nanoACQUITY UPLC系统
• Waters QTof或离子淌度质谱系统
• MSE数据采集、动态团块分辨率。
• 用于高蛋白质序列覆盖的综合肽离子计数信息，能够使您以最低的误报率识别大多数蛋白质。
• ProteinLynx GlobalSERVER™软件
• 培训和技术支持,包括在线课程

The Identity^E High Definition Proteomics System combines UPLC/MS^E data with a comprehensive peptide ion accounting informatics solution to catalog complex protein digest mixtures.

The system features:

• Kinetically-tuned protein digestion protocol to marginalize missed cleavages
• Direct nanoflow UPLC separations with rugged sub-2-μm Bridged Ethyl Hybrid (BEH) particle columns that deliver unprecedented peak capacity and retention time repeatability
• UPLC peak capacity and robustness accentuates the specificity of the Identity^E System’s retention time prediction model, allowing the amino acid composition of peptides, identified from matching MS^E spectra, to be corroborated
• High resolution mass spectrometry enables peptide detection using dynamic mass resolution and bandwidth to ensure consistent over-sampling of complex protein digests
• Identity^E is extendable through the Expression^E System to label-free relative and absolute quantification studies for protein biomarker discovery

The Identity^E system dramatically increases sequence coverage in this study of a digested cytosolic fraction from an E.coli lysate (three technical replicas).

Data Acquisition by Alternating Spectra Energy

UPLC/MS^E is a unique, patented technology that has an extremely high duty cycle, allowing the acquisition of high quality accurate mass data in a simple, unbiased manner.

UPLC/MS^E acquires data with two rapidly alternating MS functions. The first contains exclusively low-energy mass spectra, the second is composed of mass spectra acquired at elevated collision energy (designated as MS^E). The resulting data set contains a comprehensive time-resolved record of all detectible precursor and product ions.

Precursor and product ions are associated by both retention time alignment and peak shape, enabling even chimeric peptides and overlapping ion clusters to be successfully handled. The resulting time-aligned precursor and product ion mass lists are used for peptide identification with the Identity^E System’s peptide ion informatics.

Peptide Ion Informatics

This bioinformatics software visualizes peptides and proteins with a new multi-layered physicochemical model of protein primary structure. Peptide ion accounting informatics adapts to each new UPLC/MS^E data set using a pre-assessment survey. 250 identified peptides, per run, are used to fine-tune many of the 14 models used for both peptide and protein authentication.

After pre-assessment, an iterative database search strategy is initiated, identifying the most abundant protein within a sample by matching peptides, modified peptides, and retention times. An estimate of the absolute concentration of each protein is made by reference to an internal (protein) standard, allowing the fractions of all theoretical tryptic peptides that are plausibly detectable to be predicted. Proteins identified with fewer than the predicted number of peptides are sequence mapped for evidence of C/N-terminal processing.

When the most abundant protein is conclusively identified, all assigned peptide ions are subtracted from the working image of the data.

Multi-layered Physicochemical Model

The Identity^E System features Waters peptide ion accounting informatics. This technology visualizes peptides and proteins with a multi-layered physicochemical model of protein primary structure, developed from a post-genomic understanding of protein characteristics – an advantage unavailable in any other protein identification tool and unique to the Identity^E system.

Pre-assessment survey

Peptide ion accounting informatics adapts to each new UPLC/MS^E data set utilizing a pre-assessment survey. 250 unambiguously identified (abundant) peptides, per run, are used to fine-tune many of the 14 models used for both peptide and protein authentication. Automatic adjustment of these models on an injection-by-injection basis provides the means for the software to handle changes in chromatographic conditions (buffers and columns), enzymatic specificity, and sample complexity.

Fourteen different physicochemical attributes are automatically considered to maximize peptide identification confidence. Other conventional protein identification systems utilize only a small number of these characteristics.

Iterative Protein Identification

When the most abundant protein is conclusively identified, all assigned peptide ions are subtracted from the working image of the data. The process is then repeated for the next most abundant protein, as many times as is necessary to account for as many ions as possible within the UPLC/MS^E data set. As more proteins are identified, and their associated peptide ions are removed, the working image of the data becomes less complex – revealing proteins of decreasing abundance for identification.

Utilizing the 14 physicochemical models of the peptide ion accounting informatics, proteins are identified in order of their abundance, thus increasing the selectivity, specificity, and sensitivity of each iterative protein identification. The above data illustrates that proteins are being identified and their respective ions depleted by their abundance.

More Peptides per Protein

The Identity^E System identifies more peptides per protein, providing both higher sequence coverage and rigor to the protein ID. Over 95% (389) of the proteins were identified with greater than three peptides to match, compared to the LTQ Orbitrap and the LTQ with only 65% (244) and 33% (88), respectively.