RT Journal Article
SR Electronic
T1 Serum proteomics expands on identifying more high-affinity antibodies in immunized rabbits than deep B-cell repertoire sequencing alone
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 833871
DO 10.1101/833871
A1 Stefano Bonissonne
A1 Natalie Castellana
A1 Thiago Lima
A1 Katherine Harris
A1 Laura Davison
A1 Brian Avanzino
A1 Nathan Trinklein
A1 Anand Patel
YR 2019
UL http://biorxiv.org/content/early/2019/11/07/833871.abstract
AB Rabbits are a model for immunology studies, and monoclonal antibodies developed from rabbits have been highly sought after to empower immunoassays in a variety of other applications. High-throughput characterization of circulating serum antibodies in response to specific antigens is highly impactful for both immunology studies and antibody development. A combination of high throughput sequencing of antibody transcripts from B cells and proteomic analysis of serum antibodies, referred to as immunoproteogenomics, is applied to profile the immune response of rabbits to β-galactosidase (Beta-gal) in both recombinant antigen and peptide antigen immunization formats. The use of recombinant antigen resulted in observing 56.3% more heavy chains clones in serum than immunization with peptide antigens. Additionally, sampling peripheral blood mononuclear cells (PBMCs) for B-cell repertoire sequencing at different time points throughout the immunization was found to capture 47.8%-72.8% of total proteomically observed heavy chain clones, and would serve well in replacing sequencing the B cell rich, but more difficult to access spleen or bone marrow compartments. Despite B-cell repertoire sequencing to depths of 2M to 10M reads, we found proteomic evidence supporting at least 10% of serum antibodies are still missed. Further improvements to proteomic analysis techniques would enable more precise characterization of antibodies circulating in serum and better estimates of antibodies missed by repertoire sequencing.