title: Pre-Analytical Parameters Affecting Vascular Endothelial Growth Factor Measurement in Plasma: Identifying Confounders.
abstract: BACKGROUND
  Vascular endothelial growth factor-A (VEGF-A) is intensively investigated in various medical fields. However, comparing VEGF-A measurements is difficult because sample acquisition and pre-analytic procedures differ between studies. We therefore investigated which variables act as confounders of VEGF-A measurements.
  
  
  METHODS
  Following a standardized protocol, blood was taken at three clinical sites from six healthy participants (one male and one female participant at each center) twice one week apart. The following pre-analytical parameters were varied in order to analyze their impact on VEGF-A measurements: analyzing center, anticoagulant (EDTA vs. PECT / CTAD), cannula (butterfly vs. neonatal), type of centrifuge (swing-out vs. fixed-angle), time before and after centrifugation, filling level (completely filled vs. half-filled tubes) and analyzing method (ELISA vs. multiplex bead array). Additionally, intrapersonal variations over time and sex differences were explored. Statistical analysis was performed using a linear regression model.
  
  
  RESULTS
  The following parameters were identified as statistically significant independent confounders of VEGF-A measurements: analyzing center, anticoagulant, centrifuge, analyzing method and sex of the proband. The following parameters were no significant confounders in our data set: intrapersonal variation over one week, cannula, time before and after centrifugation and filling level of collection tubes.
  
  
  CONCLUSIONS
  VEGF-A measurement results can be affected significantly by the identified pre-analytical parameters. We recommend the use of CTAD anticoagulant, a standardized type of centrifuge and one central laboratory using the same analyzing method for all samples.
authors.affiliation: Eye Center, University of Freiburg, Freiburg, Germany.|Department of Pharmacology and Toxicology, University of Regensburg, Regensburg, Germany.
authors.forename: Johanna M
authors.initials: JM
authors.lastname: Walz
authors.affiliation: Eye Center, University of Freiburg, Freiburg, Germany.
authors.forename: Daniel
authors.initials: D
authors.lastname: Boehringer
authors.affiliation: Department of Ophthalmology, University of Ulm, Ulm, Germany.
authors.forename: Heidrun L
authors.initials: HL
authors.lastname: Deissler
authors.affiliation: Department of Pharmacology and Toxicology, University of Regensburg, Regensburg, Germany.|Novartis Pharma AG, Nuremberg, Germany.
authors.forename: Lothar
authors.initials: L
authors.lastname: Faerber
authors.affiliation: NMI Natural and Medical Sciences Institute at the University of Tuebingen, Reutlingen, Germany.
authors.forename: Jens C
authors.initials: JC
authors.lastname: Goepfert
authors.affiliation: Department of Ophthalmology, University of Muenster, Muenster, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Heiduschka
authors.affiliation: Department of Ophthalmology, University of Bonn, Bonn, Germany.
authors.forename: Susannah M
authors.initials: SM
authors.lastname: Kleeberger
authors.affiliation: Department of Ophthalmology, University of Kiel, University medical center, Kiel, Germany.
authors.forename: Alexa
authors.initials: A
authors.lastname: Klettner
authors.affiliation: Department of Ophthalmology, University of Bonn, Bonn, Germany.
authors.forename: Tim U
authors.initials: TU
authors.lastname: Krohne
authors.affiliation: NMI Natural and Medical Sciences Institute at the University of Tuebingen, Reutlingen, Germany.
authors.forename: Nicole
authors.initials: N
authors.lastname: Schneiderhan-Marra
authors.affiliation: Department for Ophthalmology, Eberhard-Karl University Tuebingen, Tuebingen, Germany.
authors.forename: Focke
authors.initials: F
authors.lastname: Ziemssen
authors.affiliation: Eye Center, University of Freiburg, Freiburg, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Stahl
mesh_terms: D000328:Adult; D001800:Blood Specimen Collection / Q000295:instrumentation* / Q000379:methods*; D002623:Chemistry Techniques, Analytical / Q000379:methods*; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D016014:Linear Models; D008297:Male; D011933:Reagent Kits, Diagnostic / Q000592:standards; D015203:Reproducibility of Results; D012737:Sex Factors; D042461:Vascular Endothelial Growth Factor A / Q000097:blood*; D055815:Young Adult
chemical_list: D011933:Reagent Kits, Diagnostic; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A