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                <term xml:lang="en">PEPTIDES</term>
                <term xml:lang="en">PHASE</term>
                <term xml:lang="en">QUANTITATION</term>
                <term xml:lang="en">MRM</term>
                <term xml:lang="en">PHOTO-SRM</term>
                <term xml:lang="en">HUMAN PLASMA</term>
                <term xml:lang="en">BIOMARKER VALIDATION</term>
                <term xml:lang="en">IMMUNOAFFINITY DEPLETION</term>
                <term xml:lang="en">LOW-ABUNDANCE PROTEINS</term>
                <term xml:lang="en">TARGETED MASS-SPECTROMETRY</term>
                <term xml:lang="en">Quantification</term>
                <term xml:lang="en">Protein</term>
                <term xml:lang="en">Prostate specific antigen</term>
                <term xml:lang="en">Mass spectrometry</term>
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              <p>Liquid chromatography (LC) coupled with tandem mass spectrometry (MS-MS) in selected reaction monitoring mode (SRM) has become a widely used technique for the quantification of protein biomarkers in plasma and has already proven to give similar results compared to the conventional immunoassays. To improve the lack of insufficient sensitivity for quantification of low abundance protein, we propose a new two dimensional liquid chromatography (2D-LC-SRM) method for the quantitation of prostate specific antigen (PSA) in human plasma. The method centers on anion exchange cartridge between reversed-phase chromatography and hydrophilic interaction liquid chromatography (HILIC) in an on-line arrangement. The use of the anionic cartridge allows an easier online transfer of the analytes between both dimensions. Moreover, it provides an additional selectivity since the more basic peptides are not retained on this support. This setup has been applied to the quantification of prostate specific antigen (PSA) protein in plasma on a previous generation of mass spectrometer, which enabled a limit of quantification (LOQ) of 1ng/mL without any upfront immuno-depletion or intense off-line fractionation before the SRM analysis. The obtained LOQ is compatible with the required sensitivity for the clinically relevant plasma-based PSA tests.</p>
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