title: Effect of blood sampling, processing, and storage on the measurement of complement activation biomarkers.
abstract: OBJECTIVE
  Recent studies have shown that complement hyperactivation contributes to development of thrombotic microangiopathy. The evaluation of complement biomarkers is known to be influenced by inappropriate specimen handling. However, there has been no study fully addressing this topic.
  
  
  METHODS
  Blood from each donor was subjected to 62 different handling conditions prior to complement assays.
  
  
  RESULTS
  Complement biomarkers (C4d/C3a/factor Bb/C5a/C5b-9) are stable at room temperature (RT) for up to 4 hours in whole blood containing citrate or EDTA. However, under similar conditions, levels of C4d and C3a were significantly higher in serum than those in plasma. Thawing of the samples on ice or at RT had no significant effect on complement levels. In contrast, thawing at 37°C resulted in striking increases in levels of the complement system in serum and citrated plasma but not in EDTA plasma. Up to four freeze/thaw cycles on ice or RT did not substantially increase the levels of C3a, factor Bb, C5a, and C5b-9 but had a significant effect on C4d. Long-term storage of citrated plasma at -80°C for up to 6 years had no significant effect on levels of complement factors.
  
  
  CONCLUSIONS
  The results from this study thus provide crucial guidelines for future investigations using complement biomarkers to define the role of complement system in disease.
authors.affiliation: From the Department of Pathology and Internal Medicine, College of Medicine, The Ohio State University, Columbus, and.
authors.forename: Shangbin
authors.initials: S
authors.lastname: Yang
authors.affiliation: From the Department of Pathology and Internal Medicine, College of Medicine, The Ohio State University, Columbus, and.
authors.forename: Michael
authors.initials: M
authors.lastname: McGookey
authors.affiliation: Translational Medicine, Alexion Corporation, Cheshire, CT.
authors.forename: Yi
authors.initials: Y
authors.lastname: Wang
authors.affiliation: From the Department of Pathology and Internal Medicine, College of Medicine, The Ohio State University, Columbus, and.
authors.forename: Spero R
authors.initials: SR
authors.lastname: Cataland
authors.affiliation: From the Department of Pathology and Internal Medicine, College of Medicine, The Ohio State University, Columbus, and haifeng.wu@osumc.edu.
authors.forename: Haifeng M
authors.initials: HM
authors.lastname: Wu
mesh_terms: D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D001793:Blood Preservation / Q000379:methods; D001800:Blood Specimen Collection / Q000379:methods; D003167:Complement Activation; D003165:Complement System Proteins / Q000032:analysis / Q000378:metabolism*; D006801:Humans; D013048:Specimen Handling / Q000379:methods*; D013696:Temperature; D057049:Thrombotic Microangiopathies / Q000097:blood* / Q000276:immunology
chemical_list: D015415:Biomarkers; D003165:Complement System Proteins
keywords: Biomarker; Complement activation; Diagnosis; Specimen handling; Thrombotic microangiopathy