Improved detection of antibody variable region peptides through proteomics
2024
Erasmus University Medical Center, Rotterdam, Netherlands
The polyclonal repertoire of circulating antibodies potentially holds valuable information about an individual’s humoral immune state. While bottom-up proteomics is well suited for serum proteomics, the vast number of antibodies and dynamic range of serum challenge this analysis. To acquire the serum proteome more comprehensively, high-field asymmetric waveform ion-mobility spectrometry (FAIMS) or two-dimensional chromatography were incorporated into standard trypsin-based bottom-up proteomics. Thereby, the number of variable region (VR)-related spectra increased 1.7-fold with FAIMS and 10-fold with chromatography fractionation. To match antibody VRs to spectra, de novo searching and BLAST alignment were combined. Validation of this approach showed that, as peptide length increased, the de novo accuracy decreased and BLAST performance increased. Through in silico calculations on antibody repository sequences, the uniqueness of tryptic VR peptides and their suitability as antibody surrogate was determined. Approximately one-third of these peptides were unique, and about one-third of all antibodies contained at least one unique peptide.
Improved detection of tryptic immunoglobulin variable region peptides by chromatographic and gas-phase fractionation techniques
Christoph Stingl
Added on: 07-08-2024
[1] https://www.cell.com/cell-reports-methods/fulltext/S2667-2375(24)00151-6
