Parallel production and verification of protein products using a novel high-throughput screening method

School of Biotechnology, Royal Institute of Technology, AlbaNova University Center, Stockholm, Sweden.
Biotechnology Journal (Impact Factor: 3.49). 08/2011; 6(8):1018-25. DOI: 10.1002/biot.201000430
Source: PubMed


Protein production and analysis in a parallel fashion is today applied in laboratories worldwide and there is a great need to improve the techniques and systems used for this purpose. In order to save time and money, a fast and reliable screening method for analysis of protein production and also verification of the protein product is desired. Here, a micro-scale protocol for the parallel production and screening of 96 proteins in plate format is described. Protein capture was achieved using immobilized metal affinity chromatography and the product was verified using matrix-assisted laser desorption ionization time-of-flight MS. In order to obtain sufficiently high cell densities and product yield in the small-volume cultivations, the EnBase® cultivation technology was applied, which enables cultivation in as small volumes as 150 μL. Here, the efficiency of the method is demonstrated by producing 96 human, recombinant proteins, both in micro-scale and using a standard full-scale protocol and comparing the results in regard to both protein identity and sample purity. The results obtained are highly comparable to those acquired through employing standard full-scale purification protocols, thus validating this method as a successful initial screening step before protein production at a larger scale.

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Available from: Sophia Hober, Feb 24, 2015
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    • "In this way, concentration of the enzyme controls the rate at which glucose becomes available in the medium. Enhancement of recombinant protein yield in shaken microbial cultures has been demonstrated with the first generation EnBase system [1], based on a solid starch gel at the bottom of the cultivation vessel, as well as the second generation system [2-5] comprising no solid matrix but a soluble polysaccharide in the liquid medium. The second generation EnBase includes supplementation with complex nutrients, and is therefore not strictly a fed-batch with glucose as the sole available carbon source. "
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