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- Title
Optimized dual assay for the transgenes selection and screening in CHO cell line development for recombinant protein production.
- Authors
Beketova, Elena V.; Ibneeva, Liliia R.; Abdulina, Yulia A.; Dergousova, Elena A.; Filatov, Vladimir L.; Kozlovsky, Stanislav V.; Shilov, Evgeny S.; Datskevich, Petr N.; Rozov, Fedor N.
- Abstract
Objective: To develop a simple robust methodology of screening multiple CHO cell clones secreting recombinant proteins to assess their specific productivity. Results: 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. Conclusions: 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.
- Subjects
RECOMBINANT proteins; CELL lines; GENE transfection; GENETIC vectors; CLONE cells; CELL populations; CHO cell
- Publication
Biotechnology Letters, 2019, Vol 41, Issue 8/9, p929
- ISSN
0141-5492
- Publication type
Article
- DOI
10.1007/s10529-019-02711-4