Journal ArticleDOI
Rapid, continuous purification of proteins in a microfluidic device using genetically-engineered partition tags
TLDR
The microfluidic aqueous two-phase extraction forms the core component of an integrated lab-on-a-chip device comprising cell culture, lysis, purification and analysis on a single device, and does not require expensive affinity reagents or troublesome chromatographic steps.Abstract:
High-throughput screening assays of native and recombinant proteins are increasingly crucial in life science research, including fields such as drug screening and enzyme engineering. These assays are typically highly parallel, and require minute amounts of purified protein per assay. To address this need, we have developed a rapid, automated microscale process for isolating specific proteins from sub-microlitre volumes of E. Colicell lysate. Recombinant proteins are genetically tagged to drive partitioning into the PEG-rich phase of a flowing aqueous two-phase system, which removes ∼85% of contaminating proteins, as well as unwanted nucleic acids and cell debris, on a simple microfluidic device. Inclusion of the genetic tag roughly triples recovery of the autofluorescent protein AcGFP1, and also significantly improves recovery of the enzyme glutathione S-transferase (GST), from nearly zero recovery for the wild-type enzyme, up to 40% with genetic tagging. The extraction process operates continuously, with only a single step from cell lysate to purified protein, and does not require expensive affinity reagents or troublesome chromatographic steps. The two-phase system is mild and does not disrupt protein function, as evidenced by recovery of active enzymes and functional fluorescent protein from our microfluidic process. The microfluidic aqueous two-phase extraction forms the core component of an integrated lab-on-a-chip device comprising cell culture, lysis, purification and analysis on a single device.read more
Citations
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Latest Developments in Micro Total Analysis Systems
Journal ArticleDOI
Microfluidics for food, agriculture and biosystems industries
Suresh Neethirajan,Isao Kobayashi,Mitsutoshi Nakajima,Dan Wu,Saravanan Nandagopal,Francis Lin +5 more
TL;DR: This paper synthesizes information of selected microfluidics-based applications for food, agriculture and biosystems industries and provides new approaches for bioenergy research.
Journal ArticleDOI
Emerging aqueous two-phase systems: from fundamentals of interfaces to biomedical applications
Youchuang Chao,Ho Cheung Shum +1 more
TL;DR: Recent progress in understanding the dynamics at aqueous-aqueous interfaces, and in developing interface-assisted design of artificial cells and cy to-mimetic materials, fabrication of cyto- and bio-compatible microparticles, cell micropatterning, 3D bioprinting, and microfluidic separation of cells and biomolecules are summarized.
Journal ArticleDOI
Microfluidics with aqueous two-phase systems
Steffen Hardt,Thomas Hahn +1 more
TL;DR: An overview is given about research activities in which aqueous two phase systems (ATPSs) are utilized in microfluidic setups, and a range of applications has been demonstrated, extending from separation/purification schemes to the patterning of surfaces covered with cells.
Journal ArticleDOI
Monodisperse hydrogel microspheres by forced droplet formation in aqueous two-phase systems
Iwona Ziemecka,Volkert van Steijn,Ger J. M. Koper,Michel Rosso,Aurélie Brizard,Jan H. van Esch,Michiel T. Kreutzer +6 more
TL;DR: This work shows how droplet-based microfluidics can be used in all-aqueous, surfactant-free, organic-solvent-free biocompatible two-phase environment.
References
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