RELATIVE QUANTIFICATION OF STABLE ISOTOPE LABELED PEPTIDES

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目及 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 复杂生物样品的定量分析已成为蛋白质组学领域的一个重要研究方向。虽然定量蛋白质组学实验仍然具有挑战性，但这些策略已大大促进了新的高性能质谱仪的发展。在这项工作中，我们已经评估了使用LTQ-轨道阱，一种混合质谱仪，其中线性离子阱耦合到轨道阱质量分析仪，用于定量分析。通过分析一系列酵母蛋白标准品，我们发现LTQ-Orbitrap的高质量准确度，高分辨率，大离子容量和大动态范围导致与LTQ进行的类似分析相比，肽比测量的数量和质量提高了4-5倍。我们还成功地量化了大脑发育过程中代谢标记的大鼠突触中发生的蛋白质表达差异，以进一步证明LTQ-Orbitrap用于复杂组织样品比较分析的适用性。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The quantitative analysis of complex biological samples has emerged as a key research area in the field of proteomics. Although quantitative proteomic experiments remain challenging, these strategies have been greatly facilitated by the development of newer high-performance mass spectrometers. In this work, we have evaluated the use of the LTQ-Orbitrap, a hybrid mass spectrometer in which a linear ion trap is coupled to an Orbitrap mass analyzer, for quantitative analyses. By analyzing a range of yeast protein standards, we found that the high mass accuracy, high resolution, large ion capacity, and large dynamic range of the LTQ-Orbitrap led to as much as a 4-5-fold improvement in the number and quality of the peptide ratio measurements compared to similar analyses done on the LTQ. We also successfully quantified protein expression differences that occur in metabolically labeled rat synapses during brain development to further demonstrate the suitability of the LTQ-Orbitrap for the comparative analysis of complex tissue samples.