Journal of immunology (Baltimore, Md. : 1950) · 2026
Highly efficient and low-cost single-cell culture platform for unbiased analysis of human memory B cell repertoire and antibody discovery.
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Conde Luciana, Oliveira Debora L, Maciel Gabriela, Castro Fernando, de Oliveira Albuquerque Aline, Rodrigues Danielle A, Mëllo Yare, Meira de Assis Gustavo, Gabrielle Bárbara, Ferreira Suyane S, Cunha Marcela S, Navas Carlena, Emiliano Manuela C, Rocha Marcele N, Soares Barbara, Tostes Lucas, Caloba Philippe, Maia Bruno, Bastos de Oliveira Francisco M, Tanuri Amilcar, Ferreira Orlando C, Castineiras Terezinha M, Martins da Silva João Hermínio, Echevarria Juliana, T Bozza Marcelo, Castilho Leda R, da Costa Luciana J, Felicori Liza F, Nobrega Alberto, Victora Gabriel D, Lucas Carolina, Bonomo Adriana, Vale André M
Аннотация
The SARS-CoV-2 pandemic underscored the need for innovative approaches to study humoral immunity and isolate monoclonal antibodies (mAbs) with diagnostic and therapeutic potential. Current methods for repertoire analysis at the clonal level require large-scale recombinant mAb production, limiting accessibility and delaying functional insight. We developed a single-cell culture (SCC) platform that enables profiling of human memory B cells and direct recovery of functional mAbs. Using samples from COVID-19 convalescent and vaccinated donors, we optimized SCCs with NB21 feeder cells, R848, and IL-2, achieving efficient clonal expansion and antibody secretion in short-term cultures. Screening and pseudovirus neutralization assays were performed directly with culture supernatants, bypassing the need for early recombinant antibody production. Antigen-baited cytometry sorting enriched spike-specific memory B cells by ∼30-fold. Among 592 isolated mAbs, 53% bound the Wuhan spike, targeting the receptor-binding domain (28%), N-terminal domain (15%), or other regions (57%). Cross-reactivity analysis revealed that 40% of anti-spike mAbs recognized all tested variants of concern. VH/VL sequencing uncovered convergent rearrangements, including public V3-30 and V3-53/V3-66 clones, consistent with global findings. Two public receptor-binding domain-specific antibodies demonstrated broad neutralization when produced recombinantly. Together, these results validate the SCC system as a streamlined approach for unbiased repertoire analysis and functional mAb isolation. More broadly, the platform provides a practical framework for linking B cell clonal composition with antigen specificity and serum antibody responses. By reducing costs and simplifying workflows, it expands opportunities for antibody discovery and immunoepidemiological studies, fostering wider global participation in therapeutic antibody research.