PT  - JOURNAL ARTICLE
AU  - Conor McMahon
AU  - Alexander S. Baier
AU  - Sanduo Zheng
AU  - Roberta Pascolutti
AU  - Janice X. Ong
AU  - Sarah C. Erlandson
AU  - Daniel Hilger
AU  - Aaron M. Ring
AU  - Aashish Manglik
AU  - Andrew C. Kruse
TI  - Platform for rapid nanobody discovery <em>in vitro</em>
AID  - 10.1101/151043
DP  - 2017 Jan 01
TA  - bioRxiv
PG  - 151043
4099  - http://biorxiv.org/content/early/2017/06/16/151043.short
4100  - http://biorxiv.org/content/early/2017/06/16/151043.full
AB  - Camelid single-domain antibody fragments (“nanobodies”) provide the remarkable specificity of antibodies within a single immunoglobulin VHH domain. This unique feature enables applications ranging from their use as biochemical tools to therapeutic agents. Virtually all nanobodies reported to date have been obtained by animal immunization, a bottleneck restricting many applications of this technology. To solve this problem, we developed a fully in vitro platform for nanobody discovery based on yeast surface display of a synthetic nanobody scaffold. This platform provides a facile and cost-effective method for rapidly isolating nanobodies targeting a diverse range of antigens. We provide a blueprint for identifying nanobodies starting from both purified and non-purified antigens, and in addition, we demonstrate application of the platform to discover rare conformationally-selective nanobodies to a lipid flippase and a G protein-coupled receptor. To facilitate broad deployment of this platform, we have made the library and all associated protocols publicly available.