Direct surface sampling of dried blood spots coupled with mass spectrometry for haemoglobin analysis
Haemoglobinopathies are inherited disorders, typically detected during neonatal healthcare screening programmes. Haemoglobinopathies are characterised by either a reduction in the synthesis of the globin chains or by point mutations in the globin gene often leading to a single amino acid substitutio...
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University of Birmingham
2013
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| Online Access: | https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.583137 |
| Summary: | Haemoglobinopathies are inherited disorders, typically detected during neonatal healthcare screening programmes. Haemoglobinopathies are characterised by either a reduction in the synthesis of the globin chains or by point mutations in the globin gene often leading to a single amino acid substitution in the globin chain. Current screening techniques analyse samples from resolubilised dried blood spots (DBS) by HPLC and/or isoelectric focusing. These methods are characterised by lengthy sample preparation and/or ambiguous variant determination. Further analysis is required for unequivocal diagnosis. The work presented here describes a method for the unambiguous diagnosis of haemoglobin variants in neonatal DBS samples using a surface sampling technique called liquid extraction surface analysis (LESA) coupled with high resolution top-down mass spectrometry. LESA allows DBS samples to be sampled directly and the material recovered to be directly electrosprayed into a mass spectrometer. Both the sampling and MS/MS (CID/ETD) parameters were optimised to yield maximum globin chain sequence coverage (up to 81 %) allowing for the unambiguous diagnosis of common Hb variants such as HbS and so-called unknown variants (variants unable to be diagnosed during the standard screening protocol). The LESA sampling technique has also successfully been applied to study Hb non-covalent interactions. |
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