title: Sources of variation in factor VIII, von Willebrand factor and fibrinogen measurements: implications for detecting deficiencies and increased plasma levels.
abstract: OBJECTIVE
  Differences in pre-analytical and assay conditions, inappropriate reference ranges, or inflammation may have the potential to impair clinical decisions based on measurements of factor VIII (FVIII), von Willebrand factor (VWF) and fibrinogen (Fg). This study examined the impact on FVIII, VWF and Fg in plasma of freezing and thawing, different citrate anticoagulant concentrations, and inflammation, as determined by high-sensitivity C-reactive protein (hsCRP).
  
  
  METHODS
  FVIII was determined prior to freezing and after thawing using a one-stage clotting assay (FVIII:C), an amidolytic assay (FVIII:AM) and an enzyme immunoassay (FVIII:Ag). Samples were anticoagulated with 106 or 129 mmol/L of citrate. FVIII, VWF and Fg were quantified in 300 individuals to establish reference ranges and to investigate associations with hsCRP.
  
  
  RESULTS
  Freezing and thawing reduced FVIII:C and FVIII:AM markedly. FVIII coagulant activities were not significantly different between samples anticoagulated with 106 or 129 mmol/L of citrate, respectively. FVIII, VWF and Fg were significantly associated with hsCRP. FVIII:C was greater than FVIII:AM and FVIII:Ag in all experiments, indicating that the presence of activated FVIII may lead to overestimation of FVIII:C.
  
  
  CONCLUSIONS
  Standardized freezing and thawing of plasma samples appears to be indispensable if reliable FVIII results are to be obtained. Because inflammation can potentially mask deficiency states or mimic an increased risk of thrombosis, FVIII, VWF and Fg determinations should be supplemented by measurements of hsCRP.
authors.affiliation: Institute of Hemostaseology and Transfusion Medicine, Academic City Hospital, Ludwigshafen, Germany.
authors.forename: Juergen
authors.initials: J
authors.lastname: Bach
authors.forename: Hanne
authors.initials: H
authors.lastname: Haubelt
authors.forename: Peter
authors.initials: P
authors.lastname: Hellstern
mesh_terms: D000293:Adolescent; D000328:Adult; D000347:Afibrinogenemia / Q000097:blood / Q000175:diagnosis*; D000368:Aged; D000925:Anticoagulants / Q000737:chemistry; D015415:Biomarkers / Q000097:blood; D001780:Blood Coagulation Tests / Q000592:standards; D001800:Blood Specimen Collection / Q000592:standards; D002097:C-Reactive Protein / Q000032:analysis; D002951:Citrates / Q000737:chemistry; D005169:Factor VIII / Q000032:analysis*; D005260:Female; D005340:Fibrinogen / Q000032:analysis*; D005615:Freezing; D005858:Germany; D006467:Hemophilia A / Q000097:blood / Q000175:diagnosis*; D006801:Humans; D007249:Inflammation / Q000097:blood; D008297:Male; D008875:Middle Aged; D011237:Predictive Value of Tests; D055550:Protein Stability; D012015:Reference Standards; D015203:Reproducibility of Results; D055815:Young Adult; D014842:von Willebrand Diseases / Q000097:blood / Q000175:diagnosis*; D014841:von Willebrand Factor / Q000032:analysis*
chemical_list: D000925:Anticoagulants; D015415:Biomarkers; D002951:Citrates; D014841:von Willebrand Factor; D005169:Factor VIII; D005340:Fibrinogen; D002097:C-Reactive Protein