ArticleIntegrative biology : quantitative biosciences from nano to macro2015
Microfluidics-based selection of red-fluorescent proteins with decreased rates of photobleaching.
Article in Integrative biology : quantitative biosciences from nano to macro, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 14 papers.
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Who cites it
14 citing papers in PubMed.
- Dark-State-Mediated Photobleaching in mCherry-Based Red Fluorescent Proteins.The journal of physical chemistry letters · 2026Article
- Bright and photostable yellow fluorescent proteins for extended imaging.Nature communications · 2025Article
- Automating the High-Throughput Screening of Protein-Based Optical Indicators and Actuators.Biochemistry · 2023Review
- Rapid directed molecular evolution of fluorescent proteins in mammalian cells.Protein science : a publication of the Protein Society · 2022Article
- Microscopic Imaging Methods for Organ-on-a-Chip Platforms.Micromachines · 2022Review
- Versatile phenotype-activated cell sorting.Science advances · 2020Article
- Expression and Characterization of a Bright Far-red Fluorescent Protein from the Pink-Pigmented Tissues of Porites lobata.Marine biotechnology (New York, N.Y.) · 2020Article
- Directed evolution of excited state lifetime and brightness in FusionRed using a microfluidic sorter.Integrative biology : quantitative biosciences from nano to macro · 2018Article
- Enhancing fluorescent protein photostability through robot-assisted photobleaching.Integrative biology : quantitative biosciences from nano to macro · 2018Article
- A robotic multidimensional directed evolution approach applied to fluorescent voltage reporters.Nature chemical biology · 2018Article
- High-throughput, image-based screening of pooled genetic-variant libraries.Nature methods · 2017Article
- Bacterial Phytochromes, Cyanobacteriochromes and Allophycocyanins as a Source of Near-Infrared Fluorescent Probes.International journal of molecular sciences · 2017Review
- Microfluidic System for In-Flow Reversible Photoswitching of Near-Infrared Fluorescent Proteins.Analytical chemistry · 2016Article
- Flow-Based Single Cell Deposition for High-Throughput Screening of Protein Libraries.PloS one · 2015Article
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Authors and funding
8 authors.
Funding
Abstract
Fluorescent proteins offer exceptional labeling specificity in living cells and organisms. Unfortunately, their photophysical properties remain far from ideal for long-term imaging of low-abundance cellular constituents, in large part because of their poor photostability. Despite widespread engineering efforts, improving the photostability of fluorescent proteins remains challenging due to lack of appropriate high-throughput selection methods. Here, we use molecular dynamics guided mutagenesis in conjunction with a recently developed microfluidic-based platform, which sorts cells based on their fluorescence photostability, to identify red fluorescent proteins with decreased photobleaching from a HeLa cell-based library. The identified mutant, named Kriek, has 2.5- and 4-fold higher photostability than its progenitor, mCherry, under widefield and confocal illumination, respectively. Furthermore, the results provide insight into mechanisms for enhancing photostability and their connections with other photophysical processes, thereby providing direction for ongoing development of fluorescent proteins with improved single-molecule and low-copy imaging capabilities.
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