Unlocking the Secrets of SARS-CoV-2 Immunity: A Deep Dive into Serum Antibodies and B Cell Repertoires
Project Summary
We’re excited to share our latest findings on the complex interplay between serum antibodies and B cell repertoires in individuals with SARS-CoV-2 immunity. Our study offers new insights into the development of high-affinity antibodies against emerging variants and the potential implications for Long COVID.
The ongoing global health threat posed by SARS-CoV-2 and its rapidly evolving variants has necessitated a continuous effort to develop effective monoclonal antibody therapeutics. While early antibodies showed promise, the emergence of escape variants has rendered many of them ineffective against current Omicron subvariants
To address this challenge, we employed our Alicanto technology, a proteogenomics approach that combines next-generation sequencing of immunoglobulin transcripts with mass spectrometry analysis of antibody proteins. This method allowed us to deeply characterize anti-SARS-CoV-2 serum antibodies and compare them to B cell repertoires. For the full story, check out our preprint Serum proteomics reveals high-affinity and convergent antibodies by tracking SARS-CoV2 hybrid immunity to emerging variants of concern.
Methods:
Patient Screening
In this study, we focused on three donors:
-
Two who were vaccinated and later experienced Omicron breakthrough infections
-
One who was naturally infected with SARS-CoV-2 and subsequently vaccinated
All donors had mild symptoms, and we collected whole blood samples at least four months after their most recent boost or breakthrough infection. The whole blood was processed into plasma and B cells.
Serum antibody analysis
Total IgG was purified from plasma, followed by affinity purification using either recombinant Spike trimer or the receptor binding domain (RBD) of the Omicron (B.1.1.529) virus. Antigen-purified antibodies were analyzed by nano-LC-MS/MS.
B cell sequencing
B cells expressing Spike trimer-reactive B-cell receptors (BCRs) were isolated using magnetic beads coupled to antigen. Single B cell libraries were generated using the 10X Genomics Chromium to retain heavy and light chain pairing.
Alicanto analysis
The Alicanto software matched the mass spectra to the BCR repertoire and scored each antibody clone based on proteomic and genomic evidence. A set of 24 antibodies was selected to broadly sample each individual’s immune response.
Candidate testing
Selected antibodies were recombinantly expressed as IgG1. Each antibody was tested for binding to RBD, the N-terminal domain (NTD), or the Spike trimer of Omicron B.1.1.529 for coarse-grain epitope mapping. In addition, each antibody was tested against the RBD of three SARS-CoV-2 variants, wildtype, B1.1.529 and an Omicron variant that emerged after the study had begun. Finally, each antibody was tested for neutralization of RBD to angiotension-converting enzyme-2 (ACE2) receptor in vitro.

Results:

Summary of reactivity and neutralization of the tested candidates. Candidates are grouped by donor. The left table (green) shows coarse grain epitope mapping to two domains on the Spike protein, RBD and NTD. The center table (orange) shows reactivity to three SARS-CoV-2 variants, wild-type, Omicron B.1.1.529, and BQ.1.1. The right table (blue) shows in vitro neutralization of ACE2 binding to RBD.
Key Findings
-
High-Affinity Antibodies: We identified serum antibodies with subnanomolar affinity to the receptor-binding domain (RBD) of the Omicron subvariant B.1.1.529. Remarkably, these antibodies also showed binding activity to the more recent BQ.1.1 subvariant.
-
Convergent Immune Response: Our donors exhibited a convergent immune response, with serum antibodies and B cell sequences showing similarities to previously reported anti-SARS-CoV-2 antibodies.
-
Epitope Coverage: Recombinantly expressed antibodies from our panel demonstrated binding to both the RBD and N-terminal domain of the spike protein, suggesting broad epitope coverage.
-
Cross-Reactivity: We observed antibodies retaining binding activity to the BQ.1.1 variant, despite it emerging more than five months after our donors’ infections. This finding highlights the potential for broad protection against emerging variants.
-
Serum-B Cell Dynamics: Our analysis revealed that serum antibodies represent only a small subset of the anti-SARS-CoV-2 antibodies present in the B cell repertoire, underscoring the complex dynamics between B cells and circulating antibodies.
Related Posts
Case Study: Reptor Biological vs Technical Replicates
Replicates of immune repertoire sequencing projects can provide invaluable information about the total diversity of a sample.
New Service: Alicanto for Antibody Discovery from Human Serum
We released Alicanto for Antibody Discovery from Human Serum at PEGS Boston! Click to watch our CEO, Dr. Natalie Castellana’s talk!
Case Study: Reptor Phage Enrichment
This case study aimed to identify Cas9-binding single domain antibodies using phage display and Reptor immune repertoire sequencing.
Get In Touch