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Bottom up proteomics analysis of complex mixtures of proteins commonly begin with either a protein-level separation (i.e. SDS-PAGE, iso-electric focusing) or a peptide level separation (Mudpit) before the proteins are identified by LC-MS/MS. Here we will compare the efficiency of two different types of two-dimensional peptide-level separations. We will compare the total proteins identified by liquid chromatography tandem mass spectrometry, sequence coverage and dynamic range. The two techniques differ only in the 1st dimension: the first being iso-electric focusing of peptides in pH range 4.2-6.2 (Protein Forest dPC peptide chip) and the second being strong cation exchange (SCX) chromatography and timed fraction collection of peptides.