Revolutionizing Single B Cell Screening with High-Throughput Antibody Discovery

Antibody discovery technologies are crucial for the rapid development of therapeutic agents, especially in the face of emerging pathogens. Two recent studies have proposed methodologies for improving the identification and characterization of antigen-specific B cells. These approaches focus on addressing the challenges in high-throughput screening and target the discovery of therapeutic monoclonal antibodies (mAbs) through innovative techniques.

The first paper by Lomakin et al. introduces a two-dimensional high-throughput screening method combining phage display with lentiviral immunoglobulin libraries, enabling the efficient identification of pathogen-specific antibodies without requiring detailed knowledge of the target antigen. The second paper by Fischer et al. presents a microfluidics-enabled flow cytometry (FACS) method that encapsulates single antibody-secreting cells (ASCs) in a hydrogel matrix for rapid screening and sorting of pathogen-specific antibodies.  

High-Throughput Screening: Phage Display Meets Lentiviral Libraries

In the first study, the authors developed a two-dimensional screening platform that allows for the enrichment of immunoglobulins from convalescent or vaccinated individuals using a bacteriophage library.¹ The key feature of this platform is its ability to display polypeptides up to 500 amino acids long, representing fragments of viral proteins such as the SARS-CoV-2 spike protein. This phage display approach allows for the presentation of conformational epitopes, which are often missed by traditional peptide-based screening methods. 

By pairing this phage display library with lentiviral antibody libraries, the system allows for high-throughput screening of therapeutic antibodies. The method preserves the native pairing of heavy and light chains (VH-VL), which is critical for maintaining the natural binding specificity of antibodies. Moreover, this platform was successful in isolating antibodies that could neutralize multiple variants of SARS-CoV-2, including Delta and Omicron, underscoring the system’s adaptability and robustness. 

This technology addresses a significant challenge in B cell screening: the scarcity of antigen-specific B cells in the peripheral blood. Traditional methods such as FACS and peptide microarrays are effective but have limitations, especially when targeting conformational epitopes. The phage display system overcomes this by presenting larger, more complex epitopes, leading to the isolation of highly potent neutralizing antibodies.

Related: Understand Single B Cell Antibody Technology in One Read

Microfluidics-Enabled FACS: A Game-Changer for Antibody-Secreting Cells

The second study focuses on a novel microfluidics-enabled FACS approach to screen ASCs, a previously underexplored source of high-affinity antibodies.² ASCs, which include plasma cells and plasmablasts, are responsible for secreting antibodies into the bloodstream. Historically, these cells have been challenging to study due to the lack of surface immunoglobulins (Igs), which makes it difficult to sort them using traditional FACS methods. 

The proposed microfluidics system encapsulates single ASCs in a hydrogel matrix, allowing for the capture of secreted antibodies. Once encapsulated, the cells are screened using conventional FACS, which enables the rapid isolation of antigen-specific ASCs. This method bypasses the need for expensive and labor-intensive approaches like enzyme-linked immunospot (ELISpot) assays, and it significantly accelerates the antibody discovery process. The researchers demonstrated that they could screen millions of cells and isolate SARS-CoV-2-specific antibodies with high affinity in just two weeks. 

This technology not only streamlines antibody discovery but also opens new avenues for studying the secreted antibody repertoire. By targeting the ASC compartment, the method provides access to high-affinity antibodies that are often missed when focusing solely on memory B cells. The microfluidic approach is also scalable and adaptable, making it an ideal solution for high-throughput antibody screening in pandemic preparedness and therapeutic development.

Related: Microfluidic Technology in Biotechnology

Comparative Insights: ASC Screening vs. Traditional Memory B Cell Approaches

Traditionally, therapeutic mAbs have been sourced from memory B cells, which retain surface-bound immunoglobulins. These cells are easily sorted using antigen baits in FACS, allowing for the identification of antigen-specific clones. However, ASCs, despite their ability to secrete high-affinity antibodies, have been underutilized due to their lack of surface IgG expression. The microfluidics-enabled FACS system changes this dynamic by focusing on secreted antibodies rather than relying on surface markers. 

One of the significant advantages of ASC-derived antibodies is their typically higher affinity compared to those derived from memory B cells. In the microfluidic system, antibodies were isolated from ASCs in their full-length IgG format, making them more suitable for therapeutic development. The system was able to identify antibodies with subnanomolar affinities and potent neutralizing capacities against SARS-CoV-2. In comparison, traditional memory B cell-derived antibodies require additional steps to express and characterize full-length antibodies, adding time and complexity to the discovery process. 

High-Throughput Solutions for Pandemic Preparedness

Both technologies described in these studies provide scalable, high-throughput solutions for antibody discovery, making them invaluable tools for future pandemic preparedness. The two-dimensional phage display platform can quickly adapt to new pathogens, offering a method to rapidly screen for antibodies without prior knowledge of the antigen’s structure. The microfluidics-enabled FACS system, on the other hand, provides a fast, cost-effective way to screen large populations of ASCs, generating high-affinity antibodies in a matter of weeks. 

In the context of SARS-CoV-2, both methods demonstrated their ability to isolate potent neutralizing antibodies against variants of concern. The phage display system successfully identified antibodies that neutralized Delta and Omicron variants, while the microfluidics platform isolated antibodies with high binding affinities and neutralizing capacities within two weeks of screening. These rapid timelines are essential in responding to viral outbreaks, where speed is critical in developing therapeutic interventions. 

Expanding the Reach of Antibody Discovery 

Our High-throughput Fully Human Antibody Discovery Platform integrates Cyagen’s HUGO-Ab™ mice with Biointron’s AbDrop™ microdroplet-based single B cell screening. This powerful combination accelerates the discovery and development of fully human antibodies, reducing the time from target identification to therapeutic candidate to just three months. Learn more about the service here.

References:

1. Lomakin, Y. A., Ovchinnikova, L. A., Terekhov, S. S., Dzhelad, S. S., Yaroshevich, I., Mamedov, I., Smirnova, A., Grigoreva, T., Eliseev, I. E., Filimonova, I. N., Mokrushina, Y. A., Abrikosova, V., Rubtsova, M. P., Kostin, N. N., Simonova, M. A., Bobik, T. V., Aleshenko, N. L., Alekhin, A. I., Boitsov, V. M., . . . Gabibov, A. G. (2024). Two-dimensional high-throughput on-cell screening of immunoglobulins against broad antigen repertoires. Communications Biology, 7(1), 1-13. https://doi.org/10.1038/s42003-024-06500-2

2. Fischer, K., Lulla, A., So, T. Y., Raybould, M. I., Kohler, T. N., Carlos, J., Kaminski, T. S., Hughes, R., Pyeatt, G. L., Brear, P., Matheson, N. J., Deane, C. M., Hyvönen, M., Thaventhiran, J. E., & Hollfelder, F. (2024). Rapid discovery of monoclonal antibodies by microfluidics-enabled FACS of single pathogen-specific antibody-secreting cells. Nature Biotechnology, 1-11. https://doi.org/10.1038/s41587-024-02346-5