Analysis of native peptides in serum/plasma

Abstract

Serum/plasma contains thousands of different types of proteins and peptides that can provide valuable information on numerous inherent processes. However, the existence of high-abundance proteins and the large dynamic range of serum proteins/peptides make analysis of low-abundance proteins/peptides challenging. In particular, detailed analysis and identification of native peptides remain extremely difficult due to the tremendous variety of cleavage possibilities and posttranslational modifications. Therefore, widely ranging searches based on peptide identification are difficult. Herein, I describe the highly accurate and sensitive quantitative analysis of over 2,500 peptides. The strategy combined isobaric tag labeling, amine-reactive 6-plex tandem mass tag labeling, and a modified differential solubilization method for high-yield peptide extraction. We quantitatively analyzed six pooled plasma samples (three pre-surgery and three post-surgery) to discover potential candidate biomarker peptides of renal cell carcinoma. The concentrations of about 30 peptides were found to be altered following surgery. A preliminary validation study was conducted using 77 plasma samples to demonstrate the possibility that even unidentified potential candidate biomarker peptides can be verified using the dimethyl labeling method.