Assessing the Quality and Reproducibility of a Proteomic Platform for Clinical Stroke Biomarker Discovery

The aim of this study was to investigate the quality and reproducibility of mass spectra derived from a matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF MS) platform in a patient population undergoing carotid endarterectomy. Plasma samples were either digested with trypsin or left undigested, fractionated with either C18 or weak cation exchange (WCX) columns and analysed by MALDI-TOF MS. Quality of mass spectra for each method was assessed by baseline correction (lower area under the curve ratio indicating higher quality) and signal-to-noise ratio. Mean coefficient of variation (CV%) assessed reproducibility between repeated experiments and methods. Identified mass peak intensity differences were assessed for consistency across repeated experiments. Plasma from six patients was analysed. The quality of mass spectra was significantly better when derived from digested plasma fractionated by either WCX or C18 methods compared to undigested plasma fractionated by WCX (analysis of variance, p<0.05). Mean CV% for repeated experiments was 18% and 28% for WCX and C18 fractionated digested plasma, respectively. A small number of differences in mass peak intensities were consistently observed in repeated experiments. Repeated experiments are required to confidently identify non-random mass peak intensity differences as putative plasma biomarkers that merit further investigation.