In Vitro Selection of Antibodies Targeting Yersinia pestis Membrane Lipids Using Nanodisc-Based Antigen Presentation

Proteins are the most common targets for antibody discovery and vaccine development, but their sequence variability can limit the breadth of resulting antigens. Lipids represent an alternative class of antigens due to their structural conservation and roles in host–pathogen interactions. Here, we describe the development and optimization of an in vitro antibody selection workflow using lipid-containing nanodiscs as antigen presentation platforms to enable phage and yeast display selections under conditions adapted for these non-protein targets. Lipopolysaccharide (LPS) nanodiscs were first used as a model system to evaluate selection strategies, including competitive and subtractive approaches to reduce non-specific binders, yielding peptide and single-chain variable fragment (scFv) binders that were affinity matured to improve binding signals. The same approach was subsequently used to select scFv antibodies that recognize lipid nanodiscs prepared from Yersinia pestis membrane lipid extracts. These antibodies show binding to lipid nanodiscs derived from Y. pestis, with evidence of selectivity relative to control nanodiscs. Overall, this work establishes a workflow for antibody selection against lipid-containing nanodisc antigens and highlights practical considerations associated with these targets. The approach may be useful for generating affinity reagents to membrane-associated lipids, although further characterization is required to define antigen specificity and functional activity.