Simultaneous polyclonal antibody sequencing and epitope mapping by cryo electron microscopy and mass spectrometry – a perspective

• DOI: 10.7554/elife.101322.2
• Publication Date: 2025-03-18T13:26:07Z
• Authors: Douwe Schulte; Marta Šiborová; Lukas Käll; Joost Snijder
• Keywords: Polyclonal antibodies; Cryo-Electron Microscopy; Epitope mapping

Antibodies are a major component of adaptive immunity against invading pathogens. Here we explore possibilities for an analytical approach to characterize the antigen-specific antibody repertoire directly from secreted proteins in convalescent serum. This aims perform simultaneous sequencing and epitope mapping using combination single particle cryo-electron microscopy (cryoEM) bottom-up proteomics techniques based on mass spectrometry (LC-MS/MS). We evaluate performance deep-learning tool ModelAngelo determining de novo sequences reconstructed 3D volumes antibody-antigen complexes. demonstrate that while map quality is critical bottleneck, it possible sequence variable domains cryoEM reconstructions with accuracies up 80-90%. While rate errors exceeds typical levels somatic hypermutation, show ModelAngelo-derived can be used assign V-genes. provides functional guide assemble peptides LC-MS/MS data more accurately improves tolerance background polyclonal sequences. Following this proof-of-principle, discuss feasibility future directions repertoires.