Flow-Cytometric Isolation of Human Antibodies from a Nonimmune Saccharomyces cerevisiae Surface Display Library

Journal Article · Fri Feb 28 04:00:00 EST 2003 · Nature Biotechnology, 21(2):163-170

DOI:https://doi.org/10.1038/nbt785· OSTI ID:15003641

Feldhaus, Michael ^[1]; Siegel, Robert W ^[1]; Opresko, Lee ^[1]; Coleman, James R ^[1]; Feldhaus, Jane M ^[1]; Yeung, Yik A ^[2]; Cochran, Jennifer R ^[2]; Heinzelman, Peter ^[2]; Colby, David ^[2]; Swers, Jeffrey ^[2]; Graff, Christilyn ^[2]; Wiley, H Steven ^[1]; Wittrup, K D ^[2]

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1. BATTELLE (PACIFIC NW LAB)
2. Massachusetts Institute Of Tec

A nonimmune library of 109 human antibody scFv fragments has been cloned and expressed on the surface of yeast, and nanomolar-affinity scFvs routinely obtained by magnetic bead screening and flow cytometric sorting. The yeast library can be amplified 1010-fold without measurable loss of clonal diversity, enabling effectively indefinite expansion of the library. The expression, stability, and antigen binding properties of more than 50 isolated scFv clones were assessed directly on the yeast cell surface by immunofluorescent labeling and flow cytometry, obviating separate subcloning, expression, and purification steps and thereby expediting the isolation of novel affinity reagents. The ability to use multiplex library screening demonstrates the utility of this approach for high throughput antibody isolation for proteomics applications.

• Research Organization: Pacific Northwest National Laboratory (PNNL), Richland, WA (US)
• Sponsoring Organization: USDOE
• DOE Contract Number: AC06-76RL01830
• OSTI ID: 15003641
• Report Number(s): PNNL-SA-36938
• Journal Information: Nature Biotechnology, 21(2):163-170, Journal Name: Nature Biotechnology, 21(2):163-170 Journal Issue: 2 Vol. 21; ISSN 1546-1696; ISSN 1087-0156
• Country of Publication: United States
• Language: English

Related Subjects

60 APPLIED LIFE SCIENCES
AFFINITY
ANTIBODIES
ANTIGENS
PURIFICATION
SACCHAROMYCES CEREVISIAE
SORTING
STABILITY
T-cell receptor
YEASTS
b-cells
directed evolution
fragments
gene repertoire
periplasmic expression
phage display
high-affinity
proteins
selection