title: Mass spectrometry: a tool for enhanced detection of hemoglobin variants.
abstract: BACKGROUND
  More than 900 hemoglobin (Hb) variants are currently known. Common techniques used in Hb analysis are electrophoretic and chromatographic assays. In our laboratory, we routinely apply chromatographic methods. To ascertain whether Hb variants are missed with our procedures, we additionally analyzed all samples with mass spectrometry (MS).
  
  
  METHODS
  Database evaluation was performed using all entries made in the Hb variant database HbVar, and possible Hb variants were calculated based on DNA variations. During a 5-year period, we analyzed 2105 lysates with cation-exchange HPLC (PolyCAT A column) and reversed-phase HPLC and additionally with electrospray ionization or MALDI-TOF MS. Globin chains were identified by their molecular masses.
  
  
  RESULTS
  Database evaluation revealed that 43.2% of all possible Hbalpha- and beta-chain variants were found to date (considering only single-point mutations). Currently, 68.2% of the possible charge difference variants and only 28.7% of the neutral variants are found. Among 2105 Hb samples we identified 4 samples with Hb variants that were detected only with the MS method; 2 were new Hb variants (Hb Zurich-Hottingen and Hb Zurich-Langstrasse). With cation-exchange HPLC, 1 sample was found to be a beta-thalassemia and was identified by MS to be a beta-variant (Hb Malay). More common variants, such as Hb C, Hb D, and Hb E, and thalassemias could not be detected with the MS method.
  
  
  CONCLUSIONS
  Application of MS improves the sensitivity of Hb analysis. The combination of MS with electrophoretic and chromatographic methods is optimal for the detection of Hb variants.
authors.affiliation: Division of Clinical Chemistry and Biochemistry, Department of Pediatrics, University of Zurich, Steinwiesstrasse 75, Zurich, Switzerland.
authors.forename: Peter
authors.initials: P
authors.lastname: Kleinert
authors.forename: Marlis
authors.initials: M
authors.lastname: Schmid
authors.forename: Karin
authors.initials: K
authors.lastname: Zurbriggen
authors.forename: Oliver
authors.initials: O
authors.lastname: Speer
authors.forename: Markus
authors.initials: M
authors.lastname: Schmugge
authors.forename: Bernd
authors.initials: B
authors.lastname: Roschitzki
authors.forename: Silke S
authors.initials: SS
authors.lastname: Durka
authors.forename: Urs
authors.initials: U
authors.lastname: Leopold
authors.forename: Thomas
authors.initials: T
authors.lastname: Kuster
authors.forename: Claus W
authors.initials: CW
authors.lastname: Heizmann
authors.forename: Hannes
authors.initials: H
authors.lastname: Frischknecht
authors.forename: Heinz
authors.initials: H
authors.lastname: Troxler
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods; D016208:Databases, Factual; D005260:Female; D014644:Genetic Variation; D006454:Hemoglobins / Q000737:chemistry* / Q000235:genetics*; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008297:Male; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D006454:Hemoglobins