Optimized dual assay for the transgenes selection and screening in CHO cell line development for recombinant protein production

Autor: E. S. Shilov, S.V. Kozlovsky, Elena A Dergousova, Elena V Beketova, Fedor N. Rozov, Liliia R Ibneeva, Petr N. Datskevich, Vladimir L. Filatov, Yulia A Abdulina
Rok vydání: 2019
Předmět:
Zdroj: Biotechnology Letters. 41:929-939
ISSN: 1573-6776
0141-5492
DOI: 10.1007/s10529-019-02711-4
Popis: To develop a simple robust methodology of screening multiple CHO cell clones secreting recombinant proteins to assess their specific productivity. We developed a dual assay based on immunoassay measurements of a recombinant protein expression combined with staining of viable cells with resazurin. Following this approach, colonies can be simultaneously assessed for cell growth rate and for production of a recombinant protein. Combination of these two assays enables to estimate productivity of a recombinant protein per cell from the very early stages of a cell line development process (CLD) and exclude poor producers from further steps. Comparison of the dual assay with a standard CLD protocol followed by only analysis of protein expression level showed at least 10–20% increase in the amount of clones that can be included into pool of high-producers at early stages. This shortens duration of a typical CLD scheme from 23 to 19 weeks. Our method: (i) allows to include into workflow clones that demonstrate slow growth during single cell cloning but producing high amounts of a target protein, which otherwise would be lost in standard protocols of cells screening; (ii) can be applied for testing of DNA vectors for transfection and protein production; (iii) can be used for monitoring the heterogeneity of cell population and analysis of stable pools productivity.
Databáze: OpenAIRE
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