Surface analysis for proteomics via liquid extraction surface analysis mass spectrometry and liquid chromatography mass spectrometry

摘要

Liquid extraction surface analysis (LESA) is an ambient ionisation technique which allows direct analysis of surfaces coupled with mass spectrometry. LESA mass spectrometry has been used successfully to analyse small molecules, but there are a limited number of examples where the approach has been applied to protein analysis. The work presented here aims to develop novel applications of LESA mass spectrometry of proteins. LESA mass spectrometry was used to analyse intact proteins from polymeric membranes. The rationale for these experiments was the potential application to analyse proteins electroblotted following polyacrylamide gel electrophoresis, i.e. top-down proteomics, and in air monitoring. The subsequent focus was dried blood spot (DBS) analysis. An automated LESA based trypsin digestion protocol was developed and coupled with liquid chromatography tandem mass spectrometry to enable DBS proteomics. i.e., untargeted global protein identification via a bottom-up approach. Approaches for DBS proteomics (in the absence of LESA) were explored further using conventional digestion procedures coupled with protein depletion. LESA was also applied for targeted analysis of proteins from DBS, to determine variants of alpha-1-antitrypsin. Finally, native LESA mass spectrometry was developed to analyse non-covalent complexes from dried surfaces. Native LESA mass spectrometry successfully identified the haemoglobin tetramer directly from DBS.