Single B-cell sequencing in monoclonal antibody discovery

Monoclonal antibodies (mAbs) are highly desirable therapeutic biological agents for pharmaceutical and diagnostic applications, with growing demand for rapid discovery of mAbs against challenging targets. Antibody diversity plays a crucial role in adaptive immunity. mAbs are often screened and validated from antigen-specific single B cells from short-lived ex vivo cultures. Consequently, accurately identifying the full-length variable regions of antibody heavy and light chains in individual B cells remains a challenge. Over the past decade, numerous techniques have been developed to generate mAbs, including hybridoma, antibody phage display, and single-cell sequencing. Alternatively, mAbs can be produced independently of B cells by utilizing display technologies and computational platforms driven by artificial intelligence. Accordingly, various techniques have been developed to isolate and retrieve antibody-coding sequences of B cells by miniaturizing reaction volumes. Gene expression is calculated by averaging across cells using bulk analysis. However, the emergence of single-cell sequencing has revolutionized our understanding of cellular systems. This powerful technique identifies different cell types, their functions, and their interactions within the microenvironment. Recent advancements in microfluidics have facilitated the widespread adoption of single-cell screening. Through direct amplification of the VH and VL region encoding genes from individual human B cells, single-cell sequencing can provide valuable information about B-cell subtypes, transcriptional states, and antibody sequences. This approach allows gene expression detection at a single-cell resolution, greatly advancing transcriptomic studies. Moreover, compared to traditional methods, single B-cell sequencing requires fewer cells, making it an efficient and rapid method for obtaining mAbs while preserving the natural pairing of heavy and light chains. This approach presents an attractive opportunity for capturing positive hits early in the discovery process and eliminating the possibility of losing “good clones.” This chapter summarizes the various techniques available for mAb production, including their advantages and drawbacks.