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. 2023:2681:175-212.
doi: 10.1007/978-1-0716-3279-6_11.

Single-Cell B-Cell Sequencing to Generate Natively Paired scFab Yeast Surface Display Libraries

Nathaniel Pascual  1   2 , Theodore Belecciu  1   2 , Sam Schmidt  1   2 , Athar Nakisa  2   3 , Xuefei Huang  2   3 , Daniel Woldring  4   5
Affiliations

Single-Cell B-Cell Sequencing to Generate Natively Paired scFab Yeast Surface Display Libraries

Nathaniel Pascual et al. Methods Mol Biol. 2023.

Abstract

The immune cell profiling capabilities of single-cell RNA sequencing (scRNA-seq) are powerful tools that can be applied to the design of theranostic monoclonal antibodies (mAbs). Using scRNA-seq to determine natively paired B-cell receptor (BCR) sequences of immunized mice as a starting point for design, this method outlines a simplified workflow to express single-chain antibody fragments (scFabs) on the surface of yeast for high-throughput characterization and further refinement with directed evolution experiments. While not extensively detailed in this chapter, this method easily accommodates the implementation of a growing body of in silico tools that improve affinity and stability among a range of other developability criteria (e.g., solubility and immunogenicity).

Keywords: Antibodies; B-cell receptors; Mouse immunization; Single-cell sequencing; Yeast surface display (YSD); scFab library generation.

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