Parallel purification of serum peptides for mass spectrometry

抄録

Biological fluids such as blood and urine present a convenient screening and diagnostic media for clinical analysis. Serum peptides may serve as an indicator of organism progression from normal to diseased state. However, proteomic analysis of complex samples, such as serum or plasma is frequently influenced by the presence of high protein concentrations hindering peptide detection. These proteins suppress the ionization of native peptides during MALDI-TOF MS analysis. As a result, sample complexity reduction to lower the level of abundant proteins is rapidly becoming an essential first step of any peptide analysis scheme. Several pre-fractionation strategies using chromatographic absorbents have been employed to remove abundant proteins such as albumin. As an alternative to adsorption chromatography, we have treated mammalian (human, murine, & bovine) serum or plasma samples with ultrafiltration (UF) membranes to produce relatively protein free filtrates. The UF samples were then acidified and transferred to ZipPlateC18 for de-salting and additional purification and concentration of the peptide samples. The results demonstrated that the 10K MWCO membrane gave the optimal results based on significantly improved detection and resolution of serum peptides in the 800-4000m/z range. The sample complexity reduction technology described in this presentation has resulted in convenient and rapid protocols for the isolation and analysis of native low molecular weight peptides in biological fluids such as serum or plasma. The combination of these techniques for enhanced peptide analysis applications such as the elucidation of peptide patterns enables more rapid and efficient discovery and characterization of potential biomarkers.