Chromophore pre-maturation for improved speed and sensitivity of split-GFP monitoring of protein secretion

Hansen, Henning Gram (author): Tech Univ Denmark, Novo Nordisk Fdn Ctr Biosustainabil, Lyngby, Denmark.

von Otter, Eric (author): Nanyang Technol Univ, Sch Biol Sci, Singapore 637551, Singapore.

Uhlén, Mathias (author): KTH,Science for Life Laboratory, SciLifeLab,Proteinvetenskap,The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kongens Lyngby, Denmark

(creator_code:org_t)

Related links:: https://www.nature.c...; show more...; https://urn.kb.se/re...; https://doi.org/10.1...; show less...

Abstract Subject headings

Complementation-dependent fluorescence is a powerful way to study co-localization or interactions between biomolecules. A split-GFP variant, involving the self-associating GFP 1-10 and GFP 11, has previously provided a convenient approach to measure recombinant protein titers in cell supernatants. A limitation of this approach is the slow chromophore formation after complementation. Here, we alleviate this lag in signal generation by allowing the GFP 1-10 chromophore to mature on a solid support containing GFP 11 before applying GFP 1-10 in analyses. The pre-maturated GFP 1-10 provided up to 150-fold faster signal generation compared to the non-maturated version. Moreover, pre-maturated GFP 1-10 significantly improved the ability of discriminating between Chinese hamster ovary (CHO) cell lines secreting GFP 11-tagged erythropoietin protein at varying rates. Its improved kinetics make the pre-maturated GFP 1-10 a suitable reporter molecule for cell biology research in general, especially for ranking individual cell lines based on secretion rates of recombinant proteins.

By the author/editor: Lundqvist, Magnu ...; Thalén, Niklas; Volk, Anna-Luisa; Hansen, Henning ...; von Otter, Eric; Nygren, Per-Åke, ...; show more...; Uhlén, Mathias; Rockberg, Johan; show less...

About the subject

MEDICAL AND HEALTH SCIENCES: MEDICAL AND HEAL ...; and Medical Biotechn ...; and Medical Biotechn ...

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