Finding diabetic nephropathy biomarkers in the plasma peptidome by high-throughput magnetic bead processing and MALDI-TOF-MS analysis

Purpose and experimental design: Diabetic nephropathy (DN) is the most common cause of end‐stage renal disease and improved biomarkers would help identify high‐risk individuals. The aim of this study was to discover candidate biomarkers for DN in the plasma peptidome in an in‐house cross‐sectional cohort (n=122) of type 1 diabetic patients diagnosed with normo‐, micro‐, and macroalbuminuria. Results: Automated, high‐throughput, and reproducible (interassay median CV: 13–14%) plasma peptide profiling protocols involving RPC18 and weak cation exchange magnetic beads on a liquid handling workstation with a MALDI‐TOF‐MS readout were successfully established. Using these protocols and a combined univariate (Kruskal–Wallis) and multivariate (independent component analysis) statistical analysis approach, ten single peptides and three multi‐peptide candidate biomarkers were found. Employment of RPC18 and weak cation exchange magnetic beads proved to be complementary. Conclusions and clinical relevance: The proteins found in this study, including C3f and apolipoprotein C‐I, represent new candidate biomarkers for DN from the plasma peptidome. The automated procedures and implementation of independent components analysis provide a fast and informative system for analyzing individual patient samples in protein biomarker discovery.