Small-Scale Secretory VHH Expression in Saccharomyces cerevisiae

Michiel M. Harmsen*, Marga van Hagen-van Setten, Peter T.J. Willemsen

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingChapterAcademicpeer-review

7 Citations (Scopus)

Abstract

After isolation of a single-domain antibody (VHH) binding to an antigen of interest, the soluble VHH is often produced in Escherichia coli. However, targeting VHH expression to the secretory pathway of Saccharomyces cerevisiae (baker's yeast) enables the secretion of correctly folded, soluble, disulfide-bonded, and N-glycosylated VHHs into the culture medium. Here, we describe the small-scale production of VHHs in baker's yeast in shaker flasks using both an episomal vector and a vector requiring genomic integration for higher VHH expression levels. This expression system results in the production of VHHs linked to the natural llama long hinge region including a single cysteine residue for partial dimerization. This format is especially suitable for the development of double antibody sandwich ELISAs by passive adsorption of unlabeled VHHs to polystyrene ELISA plates, antigen capture, and detection of the antigen of interest using a second biotinylated VHH. The procedures described here for detection of foot-and-mouth disease virus can also be applied to other antigens for which suitable VHHs are available.

Original languageEnglish
Title of host publicationSingle-Domain Antibodies
Subtitle of host publicationMethods and Protocols
EditorsGreg Hussack, Kevin A. Henry
PublisherSpringer
Pages159-179
Number of pages21
ISBN (Print)9781071620755
DOIs
Publication statusPublished - 2022

Publication series

NameMethods in molecular biology (Clifton, N.J.)
PublisherSpringer Verlag
Volume2446
ISSN (Print)1064-3745

Keywords

  • Biotinylation
  • Double antibody sandwich ELISA
  • Purification
  • Saccharomyces cerevisiae
  • Shake flask
  • Yeast transformation

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