Global stability of plasma proteomes for mass spectrometry-based analyses.
Author(s): Zimmerman LJ, Li M, Yarbrough WG, Slebos R, Liebler DC
Publication: Mol Cell Proteomics, 2012, Vol. 11(6), Page
PubMed ID: 22301387 PubMed Review Paper? No
Purpose of Paper
The purpose of this paper was to determine the effects of delayed centrifugation, freeze-thaw cycling, hemolysis, and using serum rather than plasma for proteomic analysis.
Conclusion of Paper
Generally delayed centrifugation, freeze-thaw cycling, specimen hemolysis and analysis of serum rather than plasma had no effects on peptide identification, protein group identification, percent semi-tryptic peptides, or percent methionine oxidized peptides. However, some individual proteins were found to be affected by delayed centrifugation, hemolysis, and the analysis of serum rather than plasma. Further, delayed centrifugation for 4 h at 4 degrees C resulted in reduced peptide identification in plasma compared to specimens not subjected to delayed centrifugation, and serum had a higher variability in the percentage of semi-tryptic peptides than plasma.
Studies
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Study Purpose
The purpose of this study was to determine the effects of delayed centrifugation, freeze-thaw cycling, hemolysis, and using serum rather than plasma for proteomic analysis. For each freeze-thaw cycle, the 3 plasma specimens were frozen at -80 degrees C for 24 h and thawed for 30 min at room temperature. Blood from ten individuals was used for the delayed centrifugation and plasma/serum experiments, and plasma or serum was frozen after centrifugation.
Summary of Findings:
Generally, delayed centrifugation, freeze-thaw cycling, specimen hemolysis and analysis of serum rather than plasma had no effects on peptide identification, protein group identification, percent semi-tryptic peptides, or percent methionine oxidized peptides. However, delayed centrifugation for 4 h at 4 degrees C resulted in reduced peptide identification in plasma compared to specimens not subjected to delayed centrifugation. Further, serum had a higher variability in the percentage of semi-tryptic peptides than plasma. Protein level comparisons revealed less than 2 fold increases in albumin, serotransferrin, inter-alpha inhibitor H4, immunoglobulin G, hemoglobin, and fibrinogen after delayed centrifugation, 10 fold higher fibrinogen alpha in plasma than serum, and no detection of fibrinogen beta and gamma in serum. The ladder peptides derived from fibrinogen were not produced by delayed centrifugation or freeze-thaw cycling. Only the spectral counts of hemoglobin proteins were significantly affected by hemolysis.
Biospecimens
Preservative Types
- Frozen
- None (Fresh)
Diagnoses:
- Not specified
Platform:
Analyte Technology Platform Peptide LC-MS or LC-MS/MS Protein LC-MS or LC-MS/MS Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Storage Storage duration <30 min
4 h
1 day
3 days
7 days
Storage Storage temperature 4 degrees C
23 degrees C
Storage Freeze/thaw cycling 0 cycles
1 cycle
2 cycles
3 cycles
5 cycles
10 cycles
Biospecimen Preservation Type of fixation/preservation Frozen
None (fresh)
Biospecimen Aliquots and Components Blood and blood products Plasma
Serum
Biospecimen Aliquots and Components Hemolysis Absent
Present
Biospecimen Aliquots and Components Centrifugation Centrifugation delays investigated