High-throughput Screening of Surface Displayed Gene Products

Authors: Walter G.; Konthur Z.; Lehrach H.

Source: Combinatorial Chemistry & High Throughput Screening, Volume 4, Number 2, April 2001 , pp. 193-205(13)

Publisher: Bentham Science Publishers

Abstract:

With the human genome project approaching completion, there is a growing interest in functional analysis of gene products. The characterization of large numbers of proteins, their expression patterns and in vivo localisations, demands the use of automated technology that maintains a logistic link to the encoding genes. As a complementary approach, phage display is used for recombinant protein expression and the selection of interacting (binding) molecules. Cloning of libraries in filamentous bacteriophage or phagemid vectors provides a physical link between the expressed protein and its encoding DNA sequence. High-throughput technology for automated library handling and phage display selection has been developed using picking-spotting robots and a module for pin-based magnetic particle handling. This system enables simultaneous interaction screening of libraries and the selection of binders to different target molecules at high throughput. Target molecules are either displayed on high-density filter membranes (protein filters) or tag-bound to magnetic particles and can be handled as native ligands. Binding activity is confirmed by magnetic particle ELISA in the microtitre format. The whole procedure from immobilisation of target molecules to confirmed clones of binders is automatable. Using this technology, we have selected human scFv antibody fragments against expression products of human cDNA libraries.

Keywords: Surface Displayed Gene Products; AUTOMATED LIBRARY; ROBOT PICKING; ROBOT REARRAYING; ROBOT SPOTTING; HIGH-DENSITY IMAGE CAPTURE; GENE IDENTIFICATION; PROTEIN EXPRESSION LIBRARIES; CELLULAR; SOLUBLE PROTEIN

Language: English

Document Type: Review article

DOI: http://dx.doi.org/10.2174/1386207013331228

Publication date: 2001-04-01

• Combinatorial Chemistry & High Throughput Screening publishes full length original research articles and reviews describing various topics in combinatorial chemistry (e.g. small molecules, peptide, nucleic acid or phage display libraries) and/or high throughput screening (e.g. developmental, practical or theoretical). Ancillary subjects of key importance, such as robotics and informatics, will also be covered by the journal. In these respective subject areas, Combinatorial Chemistry & High Throughput Screening is intended to function as the most comprehensive and up-to-date medium available. The journal should be of value to individuals engaged in the process of drug discoveryand development, in the settings of industry, academia or government.

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• By this author: Walter G. ;  Konthur Z. ;  Lehrach H.

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