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

Nathaniel Pascual; Theodore Belecciu; Sam Schmidt; Athar Nakisa; Xuefei Huang; Daniel Woldring

doi:10.1007/978-1-0716-3279-6_11

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

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

Authors

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

¹ Department of Chemical Engineering and Materials Science, Michigan State University, East Lansing, MI, USA.
² Institute for Quantitative Health Sciences and Engineering, Michigan State University, East Lansing, MI, USA.
³ Department of Chemistry, Michigan State University, East Lansing, MI, USA.
⁴ Department of Chemical Engineering and Materials Science, Michigan State University, East Lansing, MI, USA. woldring@msu.edu.
⁵ Institute for Quantitative Health Sciences and Engineering, Michigan State University, East Lansing, MI, USA. woldring@msu.edu.

PMID: 37405649
DOI: 10.1007/978-1-0716-3279-6_11

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.

MeSH terms

Animals
Antibodies, Monoclonal* / metabolism
B-Lymphocytes
Mice
Receptors, Antigen, B-Cell / genetics
Receptors, Antigen, B-Cell / metabolism
Saccharomyces cerevisiae* / metabolism
Single-Cell Analysis

Substances

Antibodies, Monoclonal
Receptors, Antigen, B-Cell