Three protein samples from seminomatous or non-seminomatous groups were randomly selected for the proteomic analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Samples were pooled (n = 3) using the same amount of protein from each sample and each pool was assessed as an individual sample. A Finnigan LTQ-Orbitrap Elite hybrid mass spectrometer (Thermo Fisher Scientific, Waltham, MA, USA) was used as previously described [45 (link),46 (link)]. Scaffold (Proteome Software Inc., Portland, OR, USA; version 4.0.6.1) was used for the identification of the differentially expressed proteins (DEPs) between the seminomatous and non-seminomatous groups. The spectral counts were used to determine the abundance of each protein (very low, low, medium or high). The expression profile of the identified DEPs (unique, underexpressed, or overexpressed) was based on the normalized spectral abundance factor (NSAF) ratio (Supplementary Table S1 ). Proteomic analysis was conducted in compliance with the Minimum Information about a Proteomics Experiment (MIAPE) guidelines of the Human Proteome Organization’s Proteomics Standards Initiative (HUPO-PSI) [47 (link)].
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