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Cell-free protein expression and functional assay in nanowell chip format

MPG-Autoren
http://pubman.mpdl.mpg.de/cone/persons/resource/persons50069

Angenendt,  Philipp
Dept. of Vertebrate Genomics (Head: Hans Lehrach), Max Planck Institute for Molecular Genetics, Max Planck Society;

Nyarsik,  Lajos
Max Planck Society;

http://pubman.mpdl.mpg.de/cone/persons/resource/persons50510

Szaflarski,  Witold
Ribosomes, Max Planck Institute for Molecular Genetics, Max Planck Society;

http://pubman.mpdl.mpg.de/cone/persons/resource/persons50167

Glokler,  Joern
Dept. of Vertebrate Genomics (Head: Hans Lehrach), Max Planck Institute for Molecular Genetics, Max Planck Society;

http://pubman.mpdl.mpg.de/cone/persons/resource/persons50444

Nierhaus,  Knud H.
Ribosomes, Max Planck Institute for Molecular Genetics, Max Planck Society;

http://pubman.mpdl.mpg.de/cone/persons/resource/persons50409

Lehrach,  Hans
Dept. of Vertebrate Genomics (Head: Hans Lehrach), Max Planck Institute for Molecular Genetics, Max Planck Society;

Lueking,  Angelika
Max Planck Society;

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Zitation

Angenendt, P., Nyarsik, L., Szaflarski, W., Glokler, J., Nierhaus, K. H., Lehrach, H., et al. (2004). Cell-free protein expression and functional assay in nanowell chip format. Analytical Chemistry, 76(7), 1844-1849. doi:10.1021/ac035114i S0003-2700(03)05114-X.


Zitierlink: http://hdl.handle.net/11858/00-001M-0000-0010-8876-7
Zusammenfassung
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.