title: Parallel production and verification of protein products using a novel high-throughput screening method.
abstract: 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.
authors.affiliation: School of Biotechnology, Royal Institute of Technology, AlbaNova University Center, Stockholm, Sweden.
authors.forename: Hanna
authors.initials: H
authors.lastname: Tegel
authors.forename: Louise
authors.initials: L
authors.lastname: Yderland
authors.forename: Tove
authors.initials: T
authors.lastname: Boström
authors.forename: Cecilia
authors.initials: C
authors.lastname: Eriksson
authors.forename: Kaisa
authors.initials: K
authors.lastname: Ukkonen
authors.forename: Antti
authors.initials: A
authors.lastname: Vasala
authors.forename: Peter
authors.initials: P
authors.lastname: Neubauer
authors.forename: Jenny
authors.initials: J
authors.lastname: Ottosson
authors.forename: Sophia
authors.initials: S
authors.lastname: Hober
mesh_terms: D002846:Chromatography, Affinity / Q000379:methods; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D057166:High-Throughput Screening Assays; D006801:Humans; D014176:Protein Biosynthesis; D011994:Recombinant Proteins / Q000032:analysis* / Q000096:biosynthesis*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
chemical_list: D011994:Recombinant Proteins