title: Quantification by nano liquid chromatography parallel reaction monitoring mass spectrometry of human apolipoprotein A-I, apolipoprotein B, and hemoglobin A1c in dried blood spots.
abstract: Proteomic analysis of blood proteins in dried blood spots (DBS) is gaining attention as a possible replacement for measurements in plasma/serum collected by venipuncture. We aimed to develop and provisionally validate a nanoflow LC-PRM-MS method for clinical use.
  
  
  
  We used Skyline to develop a nanoflow LC-PRM-MS method to quantify glycated hemoglobin-β, apolipoprotein A-I, and apolipoprotein B in DBS. Precision, linearity, interferences, and stability were determined and the method was used to analyze samples from 36 human volunteers. The method was compared with clinically validated measurements in paired blood collected via venipuncture.
  
  
  
  The method was relatively precise for these proteins (10-11% CV) and linear across the normal concentration ranges of these proteins. Interference from high total serum protein concentration (>8 g/dL) was noted for apolipoprotein A-I and apolipoprotein B. Proteins in DBS were stable for 14 days at temperatures below 25°C and trypsinized samples were stable for 48 h at 7°C. There was moderate correlation with clinical methods (r = 0.783-0.858) and significant bias in individual samples.
  
  
  
  Although the method had adequate precision and linearity for a biomarker, the accuracy compared with clinically validated assays raises concerns regarding the use of DBS compared with venipuncture for clinical use.
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, WA, USA.
authors.forename: Clark M
authors.initials: CM
authors.lastname: Henderson
authors.affiliation: Departments of Genome Sciences, University of Washington, Seattle, WA, USA.
authors.forename: James G
authors.initials: JG
authors.lastname: Bollinger
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, WA, USA.
authors.forename: Jessica O
authors.initials: JO
authors.lastname: Becker
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, WA, USA.
authors.forename: Jennifer M
authors.initials: JM
authors.lastname: Wallace
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, WA, USA.
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Laha
authors.affiliation: Departments of Genome Sciences, University of Washington, Seattle, WA, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: MacCoss
authors.affiliation: Department of Laboratory Medicine, University of Washington, Seattle, WA, USA.|Department of Medicine, University of Washington, Seattle, WA, USA.
authors.forename: Andrew N
authors.initials: AN
authors.lastname: Hoofnagle
mesh_terms: D016632:Apolipoprotein A-I / Q000097:blood*; D001055:Apolipoproteins B / Q000097:blood*; D002138:Calibration; D002853:Chromatography, Liquid / Q000379:methods*; D059788:Dried Blood Spot Testing / Q000379:methods*; D006442:Glycated Hemoglobin / Q000032:analysis*; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D036103:Nanotechnology / Q000379:methods*; D040901:Proteomics
chemical_list: D016632:Apolipoprotein A-I; D001055:Apolipoproteins B; D006442:Glycated Hemoglobin
keywords: Apolipoproteins; Biomarkers; Diabetes; Dried blood spots; LC-MS/MS; Method evaluation; Multiplexing; Provisional validation; Quantification