Effects of differences in pre-analytical processing on blood protein profiles determined with SWATH-MS.

The purpose of the present study was to assess the differences between the human serum and plasma proteomes, and the stability of human plasma proteins under different storage conditions following blood collection, by means of SWATH-MS analysis. When we compared plasma and serum prepared immediately after blood sampling, 95.5% of 176 quantified proteins differed by less than 1.5-fold. When we compared plasma samples prepared by centrifugation after storage of blood at room temperature for 0, 15 or 30 min, or under refrigeration at 0-5 °C for 1, 4 or 8 h, no protein showed a significant change (q < 0.05) that amounted to 1.5-fold or more, except hemoglobins. Those proteins were greatly increased in a single sample at 8 h, probably due to hemolysis. Comparison of data from the same samples indicates that the blood proteome is more stable than the blood metabolome. The present results suggest that most components of the proteome are essentially the same in plasma and serum, and are stable under the storage conditions examined in the present study. However, it may be important to pay attention to the extent of coagulation, and levels of platelet and hemolysis-related proteins. SIGNIFICANCE: Pre-analytical processing and storage conditions after blood collection are expected to influence the blood proteome. Therefore, we investigated differences in the proteome between human serum and plasma, as well as the stability of human plasma proteins under different storage conditions: at room temperature for 0-30 min, or at 0-5 °C for 1-8 h, which may reflect the clinical situation of blood collection. Proteomics analysis with SWATH-MS identified 342 proteins, and 176 proteins quantified with two or more unique peptides were compared. The levels of most components of the proteome were similar in plasma and serum, and were stable under the storage conditions examined. However, it is necessary to consider the possibility of coagulation, as this affects the levels of platelet and hemolysis-related proteins. Interestingly, the blood proteome appears to be more stable than the blood metabolome, based on previously reported metabolomics data with same samples. These data will be helpful in designing protocols for blood sampling and for blood biomarker discovery and validation.

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Declaration of Competing Interest S.O. is a full professor at Kumamoto University and is also a director of Proteomedix Frontiers. This study was not supported by the company, and its position at the company did not influence the design of the study, the collection of the data, the analysis or interpretation of the data, the decision to submit the manuscript for publication, or the writing of the manuscript and did not present any financial conflicts. The other authors declare no competing interests.