Zebrafish plasma analyzed by CE- and LC-MS/MS

Single analyses of a Zebrafish whole plasma digest were analyzed by both CE-MS/MS and LC-MS/MS. The identified peptides and their corresponding retention or migration times were used to train a 'blind' retention/migration time model using linear regression. This trained model provided us with information on the migration/retention coefficients resulting from the individual amino acids. All acquired tandem mass spectrometry data was processed in the Trans-Proteomic Pipeline (TPP) embedded in the Taverna scientific workflow manager. The raw data was converted to mzXML using compassXport 3.0 (Bruker) and searched with X!Tandem. The X!Tandem scores were then converted to pepXML, modeled and translated to probabilities for each peptide-spectral match by PeptideProphet. The X!Tandem search was here performed allowing a random error ±0.5 Da, +1 or +2 Da isotopic error, carbamidomethylation as fixed and oxidation as variable modification and the k-score plug-in.

本研究采用毛细管电泳-串联质谱（CE-MS/MS）与液相色谱-串联质谱（LC-MS/MS）两种分析平台，对斑马鱼全血浆酶解物分别进行单次检测。将鉴定获得的肽段及其对应的保留时间或迁移时间作为训练数据集，通过线性回归构建盲法保留/迁移时间模型，该模型可输出各氨基酸对应的迁移/保留系数。所有采集得到的串联质谱数据均通过嵌入于Taverna科学工作流管理器的串联蛋白质组学分析管线（Trans-Proteomic Pipeline，TPP）完成处理：原始数据经布鲁克（Bruker）公司的compassXport 3.0工具转换为mzXML格式后，使用X!Tandem进行数据库检索；随后将X!Tandem的打分结果转换为pepXML格式，再通过PeptideProphet工具对每条肽段-谱图匹配结果进行概率建模与转换。本次X!Tandem检索的参数设置如下：质量偏差随机误差为±0.5 Da，同位素质量偏差为+1 Da或+2 Da，以氨基甲酰甲基化为固定修饰、氧化为可变修饰，并启用k-score插件。