A 1D-PAGE/LC-ESI linear ion trap orbitrap MS approach for the analysis of synapse proteomes and synaptic protein complexes

Abstract

The 1D-PAGE/LC-ESI MS/MS approach is widely used for the qualitative and quantitative analysis of proteomes ranging from low to high complexity. As the first dimension of separation is based on SDS-PAGE, this method is compatible with the analysis of all classes of proteins including hydrophobic membrane proteins and proteins with extreme pI. 1D PAGE has the possibility to reduce protein complexity by cutting the sample gel lane into a large number of slices. The second dimension of separation is reverse phase C18 chromatography, which is coupled online to an ESI Linear Ion Trap Orbitrap MS. In this configuration the MS spectra are taken at high resolution in the Orbitrap, which improves the confidence of peptide identification. In parallel, the MS/MS spectra are generated by the Linear Ion Trap with high sensitivity, which allows the interrogation of larger numbers of peptides and thus yields improved protein identification in a given period of time. © 2011 Springer Science+Business Media, LLC.