Optimization and Standardization of Thermal Treatment as a Plasma Prefractionation Method for Proteomic Analysis
Figure 4
Label-free quantitative mass spectrometry with the multidimensional comparison of the untreated plasma, thermal treatment, IMDP, and CPLL. Tryptic peptides derived from three independent plasma samples of each group were identified at the peptide FDR<1% and quantified at the MS1 level by LC-QTOF. (a) The expression profiles of 489 unique peptides present in at least two out of three samples for a given prefractionation condition (details in Supplementary Table 4). (b) The heatmap of 58 unique proteins constituted from the 489 unique peptides which were found present with at least two peptides per protein (details in Supplementary Table 4). Both heatmaps at the peptide and protein levels showed significant clustering of three biological samples within the same group. (c) Venn diagram comparing the numbers of unique and shared identified peptides among four methods (details in Supplementary Table 5). (d) Pearson correlation of 12 samples based on their expression profiles. Numbers in the correlation matrix represent the correlation coefficient (r), where r=1 is a perfect relationship and r=0 shows no association between samples. (e) The unsupervised principal component analysis indicating that each prefractionation method produced unique sets of peptide components. These multidimensional data suggested that each prefractionation method provided a distinct plasma subproteome, which did not replace but was instead complementary to each other. The detailed information regarding all identified peptides, sequences, scores, FDR, and relative intensities are available in Supplementary Table 3. Abbreviations: FDR, false discovery rate; LC-QTOF, liquid chromatography coupled to Quadrupole-Time Of Flight.