Isolating and engineering human antibodies using yeast surface display.

Chao G; Lau WL; Hackel BJ; Sazinsky SL; Lippow SM; Wittrup KD

doi:10.1038/nprot.2006.94

Isolating and engineering human antibodies using yeast surface display.

Lau WL ,

Affiliations

1. Department of Chemical Engineering and Biological Engineering Division, Massachusetts Institute of Technology, 77 Massachusetts Avenue E19-563, Cambridge, Massachusetts 02139, USA.
Authors
Chao G¹
(1 author)

Nature Protocols, 01 Jan 2006, 1(2):755-768
https://doi.org/10.1038/nprot.2006.94 PMID: 17406305

Abstract

This protocol describes the process of isolating and engineering antibodies or proteins for increased affinity and stability using yeast surface display. Single-chain antibody fragments (scFvs) are first isolated from an existing nonimmune human library displayed on the yeast surface using magnetic-activated cell sorting selection followed by selection using flow cytometry. This enriched population is then mutagenized, and successive rounds of random mutagenesis and flow cytometry selection are done to attain desired scFv properties through directed evolution. Labeling strategies for weakly binding scFvs are also described, as well as procedures for characterizing and 'titrating' scFv clones displayed on yeast. The ultimate result of following this protocol is a panel of scFvs with increased stability and affinity for an antigen of interest.

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Read article at publisher's site: https://doi.org/10.1038/nprot.2006.94

References

Articles referenced by this article (31)

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Funding

Funders who supported this work.

NCI NIH HHS (2)

Grant ID: CA101830
20 publications
Grant ID: CA96504
36 publications

NIAID NIH HHS (1)

Grant ID: AI065824
4 publications

Search life-sciences literature (45,103,589 articles, preprints and more)

Isolating and engineering human antibodies using yeast surface display.

Affiliations

Authors

Abstract

Full text links

References

Isolation of anti-T cell receptor scFv mutants by yeast surface display.

Development of a human light chain variable domain (V(L)) intracellular antibody specific for the amino terminus of huntingtin via yeast surface display.

Directed evolution of an anti-carcinoembryonic antigen scFv with a 4-day monovalent dissociation half-time at 37 degrees C.

Molecular evolution of antibody affinity for sensitive detection of botulinum neurotoxin type A.

Directed evolution of antibody fragments with monovalent femtomolar antigen-binding affinity.

In vitro evolution of a T cell receptor with high affinity for peptide/MHC.

High-affinity CD25-binding IL-2 mutants potently stimulate persistent T cell growth.

Directed evolution of the epidermal growth factor receptor extracellular domain for expression in yeast.

Directed evolution for improved secretion of cancer-testis antigen NY-ESO-1 from yeast.

Yeast surface display for screening combinatorial polypeptide libraries.

Citations & impact

Impact metrics

Citations of article over time

Alternative metrics

Article citations

Human antibody polyreactivity is governed primarily by the heavy-chain complementarity-determining regions.

Multiplexed no-wash cellular imaging using BenzoTag, an evolved self-labeling protein.

Monoclonal antibodies: From magic bullet to precision weapon.

Protocol for engineering binding domains to recognize ligand-bound receptors by using yeast surface display.

Advancements in mammalian display technology for therapeutic antibody development and beyond: current landscape, challenges, and future prospects.

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NCI NIH HHS (2)

NIAID NIH HHS (1)

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Search life-sciences literature (45,103,589 articles, preprints and more)

Isolating and engineering human antibodies using yeast surface display.

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Funding

NCI NIH HHS (2)﻿

NIAID NIH HHS (1)﻿

Partnerships & funding

NCI NIH HHS (2)

NIAID NIH HHS (1)