Preanalytic influence of sample handling on SELDI-TOF serum protein profiles
Preanalytic influence of sample handling on SELDI-TOF serum protein profiles
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DOI：
10.1373/clinchem.2006.080101
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发表时间：
2007-04-01
影响因子：
9.3
通讯作者：
Menon, Usha
中科院分区：
文献类型：
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作者：
Timms, John F.;Arslan-Low, Elif;Menon, Usha
背景：用于人血清中疾病生物标志物发现的高通量蛋白质组学方法很有前景，但对结果的再现性和样品处理引入的变异性存在担忧。本研究调查了不同分析前处理方法对预分级血清表面增强激光解吸/电离飞行时间质谱 (SELDI-TOF MS) 蛋白质谱的影响。我们研究了样本运输时间较长的较旧收集是否会产生有用的蛋白质谱，并寻求为未来的临床蛋白质组学研究建立最可行的收集方法。方法：为了检查试管类型、凝血时间、运输/孵育时间、温度和储存方法对蛋白质谱的影响，我们使用 6 种不同的处理方法从 25 名健康志愿者收集血清。我们使用高通量预分级策略来生成阴离子交换组分，并使用表面增强激光解吸/电离飞行时间质谱法检查它们在 CM10、IMAC30-Cu 和 H50 阵列上的蛋白质谱。结果：在室温下长时间运输和孵育产生低质量峰，从而导致方案之间存在差异。最严格和最不严格的方法给出了最低的总体峰值方差，表明后者的蛋白水解可能已接近完成。对于在冰上运输的样品，凝血时间、储存方法或运输时间的影响很小。某些蛋白质（TTR、ApoCI 和转铁蛋白）不受处理影响，但其他蛋白质（ITIH4 和血红蛋白 β）显示出显着的变异性。结论：分析前处理变量的变化会影响血清蛋白的分布，包括提出的疾病生物标志物。如果所有样品的处理方式相同，则可以使用不太严格的方案对来自血清库的样品进行蛋白质组分析。 (c) 2007 年美国临床化学协会。
Background: High-throughput proteomic methods for disease biomarker discovery in human serum are promising, but concerns exist regarding reproducibility of results and variability introduced by sample handling. This study investigated the influence of different preanalytic handling methods on surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS) protein profiles of prefractionated serum. We investigated whether older collections with longer sample transit times yield useful protein profiles, and sought to establish the most feasible collection methods for future clinical proteomic studies.Methods: To examine the effect of tube type, clotting time, transport/incubation time, temperature, and storage method on protein profiles, we used 6 different handling methods to collect sera from 25 healthy volunteers. We used a high-throughput, prefractionation strategy to generate anion-exchange fractions and examined their protein profiles on CM10, IMAC30-Cu, and H50 arrays by using surf ace-enhanced laser desorption/ionization time-of-flight mass spectrometry.Results: Prolonged transport and incubation at room temperature generated low mass peaks, resulting in distinctions among the protocols. The most and least stringent methods gave the lowest overall peak variances, indicating that proteolysis in the latter may have been nearly complete. For samples transported on ice there was little effect of clotting time, storage method, or transit time. Certain proteins (TTR, ApoCI, and transferrin) were unaffected by handling, but others (ITIH4 and hemoglobin beta) displayed significant variability.Conclusions: Changes in preanalytical handling variables affect profiles of serum proteins, including proposed disease biomarkers. Proteomic analysis of samples from serum banks collected using less stringent protocols is applicable if all samples are handled identically. (c) 2007 American Association for Clinical Chemistry.