AFM-Chip-Assisted Workflow for Detection of Low-Abundance Proteins in Human Blood Plasma

Introduction

Human plasma is a highly informative biological matrix for proteomic research and biomarker discovery. However, comprehensive characterization of the plasma proteome remains challenging because protein concentrations span a wide dynamic range. Highly abundant plasma proteins frequently mask low-abundance components, limiting the analytical depth of conventional mass spectrometry workflows.

Methods

We applied an AFM-chip-assisted workflow for the enrichment of plasma proteins prior to LC-MS/MS analysis. The method is based on the preliminary concentration of proteins on chemically modified atomic force microscopy chip surfaces. Following protein adsorption, unbound components were removed, followed by on-chip enzymatic digestion, peptide extraction, and mass spectrometric identification. The workflow was evaluated as an additional sample preparation strategy for improving plasma proteome coverage.

Results

The AFM-chip-assisted workflow increased the depth of plasma proteome profiling by approximately 60% compared with conventional proteomic analysis. Approximately 33% of detected proteins were identified exclusively after AFM-chip enrichment. These findings indicate that surface-assisted protein concentration improves access to low-abundance plasma protein fractions and expands the detectable protein repertoire available for proteomic investigations.

Conclusion

AFM-chip-assisted plasma proteomics represents a promising sample preparation strategy for studies focused on the detection of proteins present at low concentrations in human plasma. By concentrating proteins prior to mass spectrometric analysis, the method complements conventional LC-MS/MS workflows and increases the depth of detectable plasma proteomes.

Funding

This work was supported by the Russian Science Foundation (project №26-15-00151).