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          Institute: MPI für molekulare Genetik     Collection: Department of Vertebrate Genomics     Display Documents

ID: 229494.0, MPI für molekulare Genetik / Department of Vertebrate Genomics
Cell-free protein expression and functional assay in nanowell chip format
Authors:Angenendt, Philipp; Nyarsik, Lajos; Szaflarski, Witold; Glökler, Jörn; Nierhaus, Knud H.; Lehrach, Hans; Cahill, Dolores J.; Lueking, Angelika
Date of Publication (YYYY-MM-DD):2004-04-01
Title of Journal:Analytical Chemistry
Journal Abbrev.:Anal Chem
Issue / Number:7
Start Page:1844
End Page:1849
Copyright:© 2004 American Chemical Society
Review Status:not specified
Audience:Experts Only
Abstract / Description:The expression and characterization of large protein libraries requires high-throughput tools for rapid and cost-effective expression and screening. A promising tool to meet these requirements is miniaturized high-density plates in chip format, consisting of an array of wells with submicroliter volumes. Here, we show the combination of nanowell chip technology and cell-free transcription and translation of proteins. Using piezoelectric dispensers, we transferred proteins into nanowells down to volumes of 100 nL and successfully detected fluorescence using confocal laser scanning. Moreover, we showed cell-free expression of proteins on a nanoliter scale using commercially available coupled transcription and translation systems. To reduce costs, we demonstrated the feasibility of diluting the coupled in vitro transcription and translation mix prior to expression. Additionally, we present an enzymatic inhibition assay in nanowells to anticipate further applications, such as the high-throughput screening of drug candidates or the identification of novel enzymes for biotechnology.
External Publication Status:published
Document Type:Article
Communicated by:Hans Lehrach
Affiliations:MPI für molekulare Genetik
External Affiliations:Ruhr-University Bochum, Medical Proteome Center, Universitätsstrasse 150, 44780 Bochum, Germany;
PROTAGEN AG, Emil-Figge-Strasse 76A, 44227 Dortmund, Germany; The Centre for Human Proteomics, Royal College of Surgeons in Ireland, Dublin 2, Ireland
DOI:10.1021/ac035114i S0003-2700(03)05114-X
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