ZIC-cHILIC Functionalized Magnetic Nanoparticle for Rapid and Sensitive Glycopeptide Enrichment from <1 µL Serum

Abstract

Due to their unique glycan composition and linkage, protein glycosylation plays significant roles in cellular function and is associated with various diseases. For comprehensive characterization of their extreme structural complexity occurring in >50% of human proteins, time-consuming multi-step enrichment of glycopeptides is required. Here we report zwitterionic n-dodecylphosphocholine-functionalized magnetic nanoparticles (ZIC-cHILIC@MNPs) as a highly efficient affinity nanoprobe for large-scale enrichment of glycopeptides. We demonstrate that ZIC-cHILIC@MNPs possess excellent affinity, with 80–91% specificity for glycopeptide enrichment, especially for sialylated glycopeptide (90%) from biofluid specimens. This strategy provides rapidity (~10 min) and high sensitivity (<1 μL serum) for the whole enrichment process in patient serum, likely due to the rapid separation using magnetic nanoparticles, fast reaction, and high performance of the affinity nanoprobe at nanoscale. Using this strategy, we achieved personalized profiles of patients with hepatitis B virus (HBV, n = 3) and hepatocellular carcinoma (HCC, n = 3) at the depth of >3000 glycopeptides, especially for the large-scale identification of under-explored sialylated glycopeptides. The glycoproteomics atlas also revealed the differential pattern of sialylated glycopeptides between HBV and HCC groups. The ZIC-cHILIC@MNPs could be a generic tool for advancing the glycoproteome analysis, and contribute to the screening of glycoprotein biomarkers.