Research Article

Journal of The American Society for Mass Spectrometry

, Volume 23, Issue 11, pp 1921-1930

First online:

Top-Down Proteomics and Direct Surface Sampling of Neonatal Dried Blood Spots: Diagnosis of Unknown Hemoglobin Variants

  • Rebecca L. EdwardsAffiliated withSchool of Biosciences, University of Birmingham
  • , Paul GriffithsAffiliated withClinical Chemistry Department, Birmingham Children’s Hospital NHS Trust
  • , Josephine BunchAffiliated withSchool of Chemistry, University of Birmingham
  • , Helen J. CooperAffiliated withSchool of Biosciences, University of Birmingham Email author 


We have previously shown that liquid microjunction surface sampling of dried blood spots coupled with high resolution top-down mass spectrometry may be used for screening of common hemoglobin variants HbS, HbC, and HbD. In order to test the robustness of the approach, we have applied the approach to unknown hemoglobin variants. Six neonatal dried blood spot samples that had been identified as variants, but which could not be diagnosed by current screening methods, were analyzed by direct surface sampling top-down mass spectrometry. Both collision-induced dissociation and electron transfer dissociation mass spectrometry were employed. Four of the samples were identified as β-chain variants: two were heterozygous Hb D-Iran, one was heterozygous Hb Headington, and one was heterozygous Hb J-Baltimore. The fifth sample was identified as the α-chain variant heterozygous Hb Phnom Penh. Analysis of the sixth sample suggested that it did not in fact contain a variant. Adoption of the approach in the clinic would require speed in both data collection and interpretation. To address that issue, we have compared manual data analysis with freely available data analysis software (ProsightPTM). The results demonstrate the power of top-down proteomics for hemoglobin variant analysis in newborn samples.

Key words

Top-down mass spectrometry Hemoglobin variants ETD CID Direct surface sampling Dried blood spots