An Economical High-Throughput Protocol for Multidimensional Fractionation of Proteins
Figure 9
Exemplary LC-MS/MS (MRM) chromatogram peptide peaks from analyses of four candidate proteins in 35% (v/v) acetonitrile eluted reversed-phase SPE fractions of 0.15 M potassium chloride eluted anion-exchange fractions, derived from DDF cytosolic extracts of six samples of cultured human Neuro-2A cells. Target protein trypsinolytic signature peptides and the employed MRM transitions were (a) Ubiquitin [TITLEVEPSDTIENVK+2H]2+; , (b) 60 s ribosomal subunit L30 [VCTLAIIDPGDSDIIR+2H]2+; , (c) serine-arginine-rich splicing factor 3 [NPPGFAFVEFEDPR+2H]2+; , and (d) PML protein [NMSERSAMAAVLAMR+2H]2+; . This data serves to show the high between sample extract precision with which some proteins were recovered and confirms the potential suitability of the described workflow for application to quantitative proteomic projects.
