Reversed phase high performance liquid chromatography coupled to tandem mass spectrometry via nano electrospray ionisation (RP-HPLC-ESI-MSMS) is widely used in the fields of proteomics to rapidly analyse the proteome of a given biological sample. In the ‘bottom-up’ approach, proteins are most commonly digested with the protease trypsin. The resultant peptide fragments are analysed to identify peptides (and subsequently the corresponding proteins) by matching the generated data to a protein database. In general, the analysis depends not only on the used instrument type but also on the ability of the chromatographic system to efficiently separate different peptides on a time scale prior to MS analysis. Here, the choice of stationary- and mobile phase components has a direct influence on the performance of the LCMS system. Usually, an increasing percentage of organic solvent in the mobile phase is used to elute peptides off the stationary column material. The most commonly used organic solvent is acetonitrile, due to its LC compatible properties. Nevertheless, alternatives like methanol/water have been demonstrated to show a superior performance in LCMS-based proteomics. In this study, the applicability of a new solvent system, consisting of a methanol/isopropanol/water mixture, has been investigated to evaluate the LCMS performance in comparison to the “more” classic solvent systems.