Jeremiah Tipton - New Reagents for Enhanced Liquid Chromatographic Separation and Charging of Intact Protein Ions for Electrospray Ionization Mass Spectrometry

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      Publication Details (including relevant citation   information):

      Valeja, S.G.; Tipton, J.D.; Emmett, M.R.;   Marshall, A.G. Analytical Chemistry, Technical Note, 82   (17), 7515-7519, (2010)


      Electrospray ionization produces multiply charged ions, thereby   lowering the mass-to-charge ratio for peptides and small proteins   to a range readily accessed by quadrupole ion trap, orbitrap, and   ion cyclotron resonance (ICR) mass analyzers   (m/z = 400−2000). For Fourier transform mass   analyzers (orbitrap and ICR), higher charge also improves   signal-to-noise ratio, mass resolution, and mass accuracy.   Addition of m-nitrobenzyl alcohol (m-NBA) or   sulfolane has previously been shown to increase the charge states   of proteins. Moreover, polar aprotic dimethylformamide (DMF)   improves chromatographic separation of proteolytic peptides for   mass analysis of solution-phase protein hydrogen/deuterium   exchange for improved (78−96%) sequence coverage. Here, we show   that addition of each of the various modifiers (DMF,   thiodiglycol, dimethylacetamide, dimethylsulfoxide, and   N-methylpyrrolidone) can significantly increase the   charge states of proteins up to 78 kDa. Moreover, incorporation   of the same modifiers into reversed-phase liquid chromatography   solvents improves sensitivity, charging, and chromatographic   resolution for intact proteins.

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