Emerging aqueous two-phase systems: from fundamentals of interfaces to biomedical applications.

Aqueous two-phase systems (ATPSs) have been recognized for their applications in extraction, separation, purification, and enrichment of (bio)molecules and cells. Recently, their unique ability to create aqueous-aqueous interfaces through phase separation and the characteristics of these interfaces have created new opportunities in biomedical applications. In this review, we summarize recent progress in understanding the dynamics at aqueous-aqueous interfaces, and in developing interface-assisted design of artificial cells and cyto-mimetic materials, fabrication of cyto- and bio-compatible microparticles, cell micropatterning, 3D bioprinting, and microfluidic separation of cells and biomolecules. We also discuss the challenges and perspectives to leverage the unique characteristics of ATPSs and their interfaces in broader applications.

[1]  Myeongsub Kim,et al.  High-Throughput Aqueous Two-Phase System Droplet Generation by Oil-Free Passive Microfluidics , 2018, ACS Omega.

[2]  Minoru Seki,et al.  Continuous cell partitioning using an aqueous two-phase flow system in microfluidic devices. , 2004, Biotechnology and bioengineering.

[3]  Lixiong Zhang,et al.  Complex Emulsions by Extracting Water from Homogeneous Solutions Comprised of Aqueous Three-Phase Systems. , 2017, Langmuir : the ACS journal of surfaces and colloids.

[4]  A. Mohamad,et al.  Electrohydrodynamic formation of single and double emulsions for low interfacial tension multiphase systems within microfluidics , 2019, Chemical Engineering Science.

[5]  Luoran Shang,et al.  Design of capillary microfluidics for spinning cell-laden microfibers , 2018, Nature Protocols.

[6]  Sze Yi Mak,et al.  Droplet Formation by Rupture of Vibration-Induced Interfacial Fingers. , 2017, Langmuir : the ACS journal of surfaces and colloids.

[7]  S. Mann,et al.  Preparation of Swellable Hydrogel-Containing Colloidosomes from Aqueous Two-Phase Pickering Emulsion Droplets. , 2018, Angewandte Chemie.

[8]  Robert H. Brown,et al.  Mutations in the vesicular trafficking protein annexin A11 are associated with amyotrophic lateral sclerosis , 2017, Science Translational Medicine.

[9]  Andrew J. deMello,et al.  Rapid cell extraction in aqueous two-phase microdroplet systems , 2010 .

[10]  Ho Cheung Shum,et al.  Fluctuation-induced dynamics of multiphase liquid jets with ultra-low interfacial tension. , 2012, Lab on a chip.

[11]  B. Mosadegh,et al.  Microprinted feeder cells guide embryonic stem cell fate. , 2011, Biotechnology and bioengineering.

[12]  Michael C. Kolios,et al.  Stable microfluidic flow focusing using hydrostatics. , 2017, Biomicrofluidics.

[13]  S. Retterer,et al.  Aqueous two-phase microdroplets with reversible phase transitions. , 2013, Lab on a chip.

[14]  T. Emrick,et al.  Reconfigurable Printed Liquids , 2018, Advanced materials.

[15]  Zhou Liu,et al.  Control of Particle Adsorption for Stability of Pickering Emulsions in Microfluidics. , 2018, Small.

[16]  Yang Song,et al.  All-aqueous multiphase microfluidics. , 2013, Biomicrofluidics.

[17]  B. Binks,et al.  Phase Inversion of Silica Particle-Stabilized Water-in-Water Emulsions. , 2019, Langmuir : the ACS journal of surfaces and colloids.

[18]  Mingqiang Li,et al.  Cell-laden microfluidic microgels for tissue regeneration. , 2016, Lab on a chip.

[19]  M. Pérez-Madrigal,et al.  Controlling the Size of Two-Dimensional Polymer Platelets for Water-in-Water Emulsifiers , 2017, ACS central science.

[20]  Yangcheng Lu,et al.  Phase separation of parallel laminar flow for aqueous two phase systems in branched microchannel , 2011 .

[21]  David A Weitz,et al.  Microfluidic fabrication of water-in-water (w/w) jets and emulsions. , 2012, Biomicrofluidics.

[22]  Philip C Bevilacqua,et al.  RNA catalysis through compartmentalization. , 2012, Nature chemistry.

[23]  L. Benyahia,et al.  Stabilization of Water-in-Water Emulsions by Nanorods. , 2016, ACS macro letters.

[24]  W. Li,et al.  Aqueous two-phase extraction for bovine serum albumin (BSA) with co-laminar flow in a simple coaxial capillary microfluidic device , 2014 .

[25]  Reinhard Lipowsky,et al.  Membrane nanotubes induced by aqueous phase separation and stabilized by spontaneous curvature , 2011, Proceedings of the National Academy of Sciences.

[26]  Young-Soo Song,et al.  Microextraction in a tetrabutylammonium bromide/ammonium sulfate aqueous two-phase system and electrohydrodynamic generation of a micro-droplet. , 2007, Journal of chromatography. A.

[27]  T. Meng,et al.  Enhanced enzymatic reaction by aqueous two-phase systems using parallel-laminar flow in a double Y-branched microfluidic device , 2018 .

[28]  D. Weitz,et al.  Microfluidic fabrication of microparticles for biomedical applications. , 2018, Chemical Society reviews.

[29]  Aviad Levin,et al.  Hierarchical Biomolecular Emulsions Using 3-D Microfluidics with Uniform Surface Chemistry. , 2017, Biomacromolecules.

[30]  A. Philipse,et al.  Colloidal zein particles at water-water interfaces , 2017 .

[31]  Sung‐Jin Kim,et al.  Passive droplet generation in aqueous two-phase systems with a variable-width microchannel. , 2019, Soft matter.

[32]  L. Benyahia,et al.  Water-In-Water Emulsion Gels Stabilized by Cellulose Nanocrystals. , 2018, Langmuir : the ACS journal of surfaces and colloids.

[33]  R. Sullivan,et al.  Emulsified and Liquid-Liquid Phase-Separated States of α-Pinene Secondary Organic Aerosol Determined Using Aerosol Optical Tweezers. , 2017, Environmental Science and Technology.

[34]  Jin Woong Kim,et al.  Affinity Partitioning-Induced Self-Assembly in Aqueous Two-Phase Systems: Templating for Polyelectrolyte Microcapsules. , 2016, ACS macro letters.

[35]  Shuichi Takayama,et al.  Polymeric Aqueous Biphasic Systems for Non‐Contact Cell Printing on Cells: Engineering Heterocellular Embryonic Stem Cell Niches , 2010, Advanced materials.

[36]  J. Chapel,et al.  Preparation of Template-Free Robust Yolk-Shell Gelled Particles from Controllably Evolved All-in-Water Emulsions. , 2018, Small.

[37]  Christine D. Keating,et al.  Aqueous Phase Separation as a Possible Route to Compartmentalization of Biological Molecules , 2012, Accounts of chemical research.

[38]  T. Mittag,et al.  Considerations and Challenges in Studying Liquid-Liquid Phase Separation and Biomolecular Condensates , 2019, Cell.

[39]  C. Keating,et al.  Combining Catalytic Microparticles with Droplets Formed by Phase Coexistence: Adsorption and Activity of Natural Clays at the Aqueous/Aqueous Interface , 2017, Scientific Reports.

[40]  Yang Song,et al.  All-Aqueous Electrosprayed Emulsion for Templated Fabrication of Cytocompatible Microcapsules. , 2015, ACS applied materials & interfaces.

[41]  Hossein Tavana,et al.  Interfacial Tension Effect on Cell Partition in Aqueous Two-Phase Systems. , 2015, ACS applied materials & interfaces.

[42]  B. Helms,et al.  Building Reconfigurable Devices Using Complex Liquid–Fluid Interfaces , 2019, Advanced materials.

[43]  Ho Cheung Shum,et al.  Phase‐Separation‐Induced Formation of Janus Droplets Based on Aqueous Two‐Phase Systems , 2017 .

[44]  S. Chueh,et al.  Microspheres of hydroxyapatite/reconstituted collagen as supports for osteoblast cell growth. , 1999, Biomaterials.

[45]  Daeyeon Lee,et al.  All-Aqueous Assemblies via Interfacial Complexation: Toward Artificial Cell and Microniche Development. , 2017, Langmuir : the ACS journal of surfaces and colloids.

[46]  Wei Zhang,et al.  Bio-inspired spontaneously regenerative tough hydrogels. , 2019, Angewandte Chemie.

[47]  E. Villermaux,et al.  Physics of liquid jets , 2008 .

[48]  C. Dobson,et al.  Fabrication and Characterization of Reconstituted Silk Microgels for the Storage and Release of Small Molecules. , 2019, Macromolecular rapid communications.

[49]  Steffen Hardt,et al.  Concentration and size separation of DNA samples at liquid-liquid interfaces. , 2011, Analytical chemistry.

[50]  M. Bier,et al.  Electrophoretic transport of solutes in aqueous two‐phase systems , 1990, Electrophoresis.

[51]  B. Helms,et al.  Bicontinuous structured liquids with sub-micrometre domains using nanoparticle surfactants. , 2017, Nature nanotechnology.

[52]  Moran Wang,et al.  Flexibility of inactive electrokinetic layer at charged solid-liquid interface in response to bulk ion concentration. , 2019, Journal of colloid and interface science.

[53]  H. Chang,et al.  Rapid separation of bacteriorhodopsin using a laminar-flow extraction system in a microfluidic device. , 2010, Biomicrofluidics.

[54]  C. Keating,et al.  RNA-Based Coacervates as a Model for Membraneless Organelles: Formation, Properties, and Interfacial Liposome Assembly. , 2016, Langmuir : the ACS journal of surfaces and colloids.

[55]  S. Hardt,et al.  Electrophoretic transport of biomolecules across liquid–liquid interfaces , 2011, Journal of physics. Condensed matter : an Institute of Physics journal.

[56]  L. Benyahia,et al.  Exploiting Complex Formation between Polysaccharides and Protein Microgels To Influence Particle Stabilization of W/W Emulsions. , 2018, Langmuir : the ACS journal of surfaces and colloids.

[57]  B. V. Schmidt.  Double Hydrophilic Block Copolymer Self-Assembly in Aqueous Solution , 2018 .

[58]  R. Lipowsky,et al.  Giant Vesicles Exposed to Aqueous Two‐Phase Systems: Membrane Wetting, Budding Processes, and Spontaneous Tubulation , 2017 .

[59]  Sze Yi Mak,et al.  Emergence of Droplets at the Nonequilibrium All-Aqueous Interface in a Vertical Hele-Shaw Cell. , 2018, Langmuir : the ACS journal of surfaces and colloids.

[60]  Pingan Zhu,et al.  Microfluidic generation of aqueous two-phase-system (ATPS) droplets by oil-droplet choppers. , 2017, Lab on a chip.

[61]  Marco Rito-Palomares,et al.  Cell‐based aqueous two‐phase systems for therapeutics , 2019, Journal of Chemical Technology & Biotechnology.

[62]  Ki-Hwan Nam,et al.  Continuous-Flow Fractionation of Animal Cells in Microfluidic Device Using Aqueous Two-Phase Extraction , 2005, Biomedical microdevices.

[63]  Jason K. Streit,et al.  Differentiating Left- and Right-Handed Carbon Nanotubes by DNA. , 2016, Journal of the American Chemical Society.

[64]  Guofu Zhou,et al.  Interfacial Complexation Induced Controllable Fabrication of Stable Polyelectrolyte Microcapsules Using All-Aqueous Droplet Microfluidics for Enzyme Release. , 2019, ACS applied materials & interfaces.

[65]  Self‐Orienting Hydrogel Micro‐Buckets as Novel Cell Carriers , 2018, Angewandte Chemie.

[66]  S. Mytnyk,et al.  Compartmentalizing Supramolecular Hydrogels Using Aqueous Multi-phase Systems. , 2017, Angewandte Chemie.

[67]  F. Hartl,et al.  Pathways of cellular proteostasis in aging and disease , 2018, The Journal of cell biology.

[68]  Droplet formation and shrinking in aqueous two-phase systems using a membrane emulsification method. , 2015, Biomicrofluidics.

[69]  Xiaoquan Yang,et al.  Slowing the Starch Digestion by Structural Modification through Preparing Zein/Pectin Particle Stabilized Water-in-Water Emulsion. , 2018, Journal of agricultural and food chemistry.

[70]  Daeyeon Lee,et al.  One-Step Generation of Cell-Encapsulating Compartments via Polyelectrolyte Complexation in an Aqueous Two Phase System. , 2016, ACS applied materials & interfaces.

[71]  Shuichi Takayama,et al.  Budding-like division of all-aqueous emulsion droplets modulated by networks of protein nanofibrils , 2018, Nature Communications.

[72]  Shoji Takeuchi,et al.  Metre-long cell-laden microfibres exhibit tissue morphologies and functions. , 2013, Nature materials.

[73]  K. Prather,et al.  Inside versus outside: ion redistribution in nitric acid reacted sea spray aerosol particles as determined by single particle analysis. , 2013, Journal of the American Chemical Society.

[74]  P. Albertsson,et al.  Partition of Proteins in Liquid Polymer–Polymer Two-Phase Systems , 1958, Nature.

[75]  T. Meng,et al.  Aqueous Two-Phase Droplet-Templated Colloidosomes Composed of Self-Formed Particles via Spatial Confined Biomineralization. , 2019, ACS applied materials & interfaces.

[76]  M. Kreutzer,et al.  Slow growth of the Rayleigh-Plateau instability in aqueous two phase systems. , 2012, Biomicrofluidics.

[77]  R. Lipowsky.  Response of Membranes and Vesicles to Capillary Forces Arising from Aqueous Two-Phase Systems and Water-in-Water Droplets. , 2018, The journal of physical chemistry. B.

[78]  Yang Song,et al.  Monodisperse w/w/w double emulsion induced by phase separation. , 2012, Langmuir : the ACS journal of surfaces and colloids.

[79]  Christine D. Keating,et al.  Biocatalyzed mineralization in an aqueous two-phase system: effect of background polymers and enzyme partitioning. , 2013, Journal of materials chemistry. B.

[80]  Christine D. Keating,et al.  Aqueous Emulsion Droplets Stabilized by Lipid Vesicles as Microcompartments for Biomimetic Mineralization. , 2015, Langmuir : the ACS journal of surfaces and colloids.

[81]  L. D. Silva,et al.  Liquid–liquid extraction of metal ions without use of organic solvent , 2008 .

[82]  J. Coutinho,et al.  A Triple Salting-Out Effect is Required for the Formation of Ionic-Liquid-Based Aqueous Multiphase Systems. , 2017, Angewandte Chemie.

[83]  T. Cubaud,et al.  Viscous Wave Breaking and Ligament Formation in Microfluidic Systems. , 2018, Physical review letters.

[84]  Ho Cheung Shum,et al.  Generation of High-Order All-Aqueous Emulsion Drops by Osmosis-Driven Phase Separation. , 2018, Small.

[85]  Trapping and assembly of living colloids at water-water interfaces. , 2014, Soft matter.

[86]  S. Takayama,et al.  Precisely targeted delivery of cells and biomolecules within microchannels using aqueous two-phase systems , 2011, Biomedical microdevices.

[87]  J. Kutter,et al.  Electrophoretic partitioning of proteins in two-phase microflows. , 2007, Lab on a chip.

[88]  Bifeng Liu,et al.  Rapid, highly efficient extraction and purification of membrane proteins using a microfluidic continuous-flow based aqueous two-phase system. , 2011, Journal of chromatography. A.

[89]  C. Brangwynne,et al.  Liquid phase condensation in cell physiology and disease , 2017, Science.

[90]  C. Keating,et al.  Budding and asymmetric protein microcompartmentation in giant vesicles containing two aqueous phases. , 2008, Journal of the American Chemical Society.

[91]  A. Hyman,et al.  Liquid-liquid phase separation in biology. , 2014, Annual review of cell and developmental biology.

[92]  C. Keating,et al.  Dynamic microcompartmentation in synthetic cells , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[93]  Ho Cheung Shum,et al.  Partitioning-dependent conversion of polyelectrolyte assemblies in an aqueous two-phase system. , 2018, Soft matter.

[94]  Daeyeon Lee,et al.  Continuous Fabrication of Hierarchical and Asymmetric Bijel Microparticles, Fibers, and Membranes by Solvent Transfer‐Induced Phase Separation (STRIPS) , 2015, Advanced materials.

[95]  Brendan M Leung,et al.  Aqueous two-phase system-derived biofilms for bacterial interaction studies. , 2012, Biomacromolecules.

[96]  Reinhard Lipowsky,et al.  Wetting-Induced Budding of Vesicles in Contact with Several Aqueous Phases , 2012, The journal of physical chemistry. B.

[97]  Yang Song,et al.  Manipulation of viscous all-aqueous jets by electrical charging. , 2013, Chemical communications.

[98]  Bradley A. Rogers,et al.  A stepwise mechanism for aqueous two-phase system formation in concentrated antibody solutions , 2019, Proceedings of the National Academy of Sciences.

[99]  H. Shum,et al.  Development of dual-component protein microparticles in all-aqueous systems for biomedical applications , 2019, Journal of Materials Chemistry B.

[100]  R. Lipowsky,et al.  Transition from complete to partial wetting within membrane compartments. , 2008, Journal of the American Chemical Society.

[101]  R. Lipowsky,et al.  Droplet-induced budding transitions of membranes , 2011, 1106.5145.

[102]  Dominique Durand,et al.  Particles trapped at the droplet interface in water-in-water emulsions. , 2012, Langmuir : the ACS journal of surfaces and colloids.

[103]  Hossein Tavana,et al.  High Throughput, Polymeric Aqueous Two‐Phase Printing of Tumor Spheroids , 2014, Advanced functional materials.

[104]  Stephen Mann,et al.  Designs for life: protocell models in the laboratory. , 2012, Chemical Society reviews.

[105]  Shabir Hassan,et al.  Aqueous Two‐Phase Emulsion Bioink‐Enabled 3D Bioprinting of Porous Hydrogels , 2018, Advanced materials.

[106]  H. Shum,et al.  All-Aqueous Thin-Film-Flow-Induced Cell-Based Monolayers. , 2019, ACS applied materials & interfaces.

[107]  V. Gnyawali,et al.  Microfluidic diamagnetic water-in-water droplets: a biocompatible cell encapsulation and manipulation platform. , 2018, Lab on a chip.

[108]  Charles R. Mace,et al.  Experimental and Theoretical Validation of System Variables That Control the Position of Particles at the Interface of Immiscible Liquids. , 2018, Langmuir : the ACS journal of surfaces and colloids.

[109]  Christine D. Keating,et al.  Liquid–liquid phase separation in artificial cells , 2018, Interface Focus.

[110]  Sung-Jin Kim,et al.  Water-head-driven microfluidic oscillators for autonomous control of periodic flows and generation of aqueous two-phase system droplets. , 2017, Lab on a chip.

[111]  P Novo,et al.  On-chip sample preparation and analyte quantification using a microfluidic aqueous two-phase extraction coupled with an immunoassay. , 2014, Lab on a chip.

[112]  Håkan Wennerström,et al.  The nature of phase separation in aqueous two-polymer systems , 1986 .

[113]  Dae Kun Hwang,et al.  Water-in-Water Droplets by Passive Microfluidic Flow Focusing. , 2016, Analytical chemistry.

[114]  Pingan Zhu,et al.  Passive and active droplet generation with microfluidics: a review. , 2016, Lab on a chip.

[115]  E. M. Blokhuis,et al.  Tension, Rigidity, and Preferential Curvature of Interfaces between Coexisting Polymer Solutions , 2013, 1304.6829.

[116]  Bagrat Grigoryan,et al.  Multivascular networks and functional intravascular topologies within biocompatible hydrogels , 2019, Science.

[117]  J. Kang,et al.  Nanopore Sensing in Aqueous Two-Phase System: Simultaneous Enhancement of Signal and Translocation Time via Conformal Coating. , 2017, Small.

[118]  S. Parekh,et al.  Microstructured Elastomer‐PEG Hydrogels via Kinetic Capture of Aqueous Liquid–Liquid Phase Separation , 2018, Advanced science.

[119]  J. V. van Duijneveldt,et al.  Stabilisation of water-in-water emulsions by montmorillonite platelets. , 2017, Journal of colloid and interface science.

[120]  B. Murray,et al.  Effect of oil droplets and their solid/liquid composition on the phase separation of protein-polysaccharide mixtures. , 2013, Langmuir : the ACS journal of surfaces and colloids.

[121]  E. Dickinson,et al.  Interfacial structuring in a phase-separating mixed biopolymer solution containing colloidal particles. , 2009, Langmuir : the ACS journal of surfaces and colloids.

[122]  G. Walker,et al.  Microfluidic aqueous two phase system for leukocyte concentration from whole blood , 2009, Biomedical microdevices.

[123]  S. Tsai,et al.  Controlled generation of spiky microparticles by ionic cross-linking within an aqueous two-phase system. , 2019, Soft matter.

[124]  Ho Cheung Shum,et al.  Syringe-pump-induced fluctuation in all-aqueous microfluidic system implications for flow rate accuracy. , 2014, Lab on a chip.

[125]  Charles R. Mace,et al.  Aqueous multiphase systems of polymers and surfactants provide self-assembling step-gradients in density. , 2012, Journal of the American Chemical Society.

[126]  Daeyeon Lee,et al.  Tuning interfacial complexation in aqueous two phase systems with polyelectrolytes and nanoparticles for compound all water emulsion bodies (AWE-somes). , 2017, Physical chemistry chemical physics : PCCP.

[127]  A. Poortinga.  Microcapsules from self-assembled colloidal particles using aqueous phase-separated polymer solutions. , 2008, Langmuir : the ACS journal of surfaces and colloids.

[128]  G. Luo,et al.  Microfluidic fabrication of water-in-water droplets encapsulated in hydrogel microfibers , 2019, Chinese Chemical Letters.

[129]  Steffen Hardt,et al.  Microfluidics with aqueous two-phase systems. , 2012, Lab on a chip.

[130]  B. Binks.  Particles as surfactants—similarities and differences , 2002 .

[131]  V. N. Paunov,et al.  Fabrication of Human Keratinocyte Cell Clusters for Skin Graft Applications by Templating Water-in-Water Pickering Emulsions , 2019, Biomimetics.

[132]  Kazuki Akamatsu,et al.  Aqueous two-phase system formation in small droplets by SPG membrane emulsification followed by water extraction. , 2019, Langmuir : the ACS journal of surfaces and colloids.

[133]  V. Gnyawali,et al.  Microneedle-assisted microfluidic flow focusing for versatile and high throughput water-in-water droplet generation. , 2019, Journal of colloid and interface science.

[134]  B. Erné,et al.  Chemical physics of water-water interfaces. , 2016, Biointerphases.

[135]  M. Dehghani,et al.  Application of polyethylene glycol based aqueous two-phase systems for extraction of heavy metals , 2017 .

[136]  B. Murray,et al.  The influence of oil droplets on the phase separation of protein–polysaccharide mixtures , 2014 .

[137]  Marco Y. Hein,et al.  A Liquid-to-Solid Phase Transition of the ALS Protein FUS Accelerated by Disease Mutation , 2015, Cell.

[138]  Yee Koon Yau,et al.  Current applications of different type of aqueous two-phase systems , 2015, Bioresources and Bioprocessing.

[139]  Y. H. Zhang,et al.  Liquid-liquid phase separation in mixed organic/inorganic single aqueous aerosol droplets. , 2015, Journal of Physical Chemistry A.

[140]  Myeongsub Kim,et al.  Flow regime mapping of aqueous two-phase system droplets in flow-focusing geometries , 2017 .

[141]  T. Ono,et al.  Microfluidic Formation of Hydrogel Microcapsules with a Single Aqueous Core by Spontaneous Cross-Linking in Aqueous Two-Phase System Droplets. , 2019, Langmuir : the ACS journal of surfaces and colloids.

[142]  Hai-Tao Liu,et al.  A Microfluidic Strategy for Controllable Generation of Water-in-Water Droplets as Biocompatible Microcarriers. , 2018, Small.

[143]  H. Daigle,et al.  On the shear stability of water-in-water Pickering emulsions stabilized with silica nanoparticles. , 2018, Journal of colloid and interface science.

[144]  Ho Cheung Shum,et al.  Musical Interfaces: Visualization and Reconstruction of Music with a Microfluidic Two-Phase Flow , 2014, Scientific Reports.

[145]  Yang Song,et al.  Fabrication of fibrillosomes from droplets stabilized by protein nanofibrils at all-aqueous interfaces , 2016, Nature Communications.

[146]  Sze Yi Mak,et al.  The dripping-to-jetting transition in a co-axial flow of aqueous two-phase systems with low interfacial tension , 2017 .

[147]  L. Sagis,et al.  Bending Rigidity of Interfaces in Aqueous Phase-Separated Biopolymer Mixtures , 2004 .

[148]  D. Weitz,et al.  Geometrically mediated breakup of drops in microfluidic devices. , 2003, Physical review letters.

[149]  B. Murray,et al.  The effect of nanoparticles on the phase separation of waxy corn starch+locust bean gum or guar gum , 2014 .

[150]  L. Leon,et al.  Oligonucleotide-Peptide Complexes: Phase Control by Hybridization. , 2018, Journal of the American Chemical Society.

[151]  S. Hardt,et al.  Size-dependent detachment of DNA molecules from liquid-liquid interfaces , 2011 .

[152]  V Chu,et al.  Design of a microfluidic platform for monoclonal antibody extraction using an aqueous two-phase system. , 2012, Journal of chromatography. A.

[153]  Peter Tompa,et al.  Polymer physics of intracellular phase transitions , 2015, Nature Physics.

[154]  V Chu,et al.  Determination of aqueous two phase system binodal curves using a microfluidic device. , 2014, Journal of chromatography. A.

[155]  Michiel T. Kreutzer,et al.  Microcapsules with a permeable hydrogel shell and an aqueous core continuously produced in a 3D microdevice by all-aqueous microfluidics , 2017 .

[156]  Ali Khademhosseini,et al.  3D Bioprinting for Tissue and Organ Fabrication , 2016, Annals of Biomedical Engineering.

[157]  E. Dickinson.  Particle-based stabilization of water-in-water emulsions containing mixed biopolymers , 2019, Trends in Food Science & Technology.

[158]  Dae Kun Hwang,et al.  Microfluidic generation of aqueous two-phase system (ATPS) droplets by controlled pulsating inlet pressures. , 2015, Lab on a chip.

[159]  Zhou Liu,et al.  Freeform, Reconfigurable Embedded Printing of All‐Aqueous 3D Architectures , 2019, Advanced materials.

[160]  S. Takayama,et al.  Rehydration of Polymeric, Aqueous, Biphasic System Facilitates High Throughput Cell Exclusion Patterning for Cell Migration Studies , 2011 .

[161]  K. Yoshikawa,et al.  Aqueous/Aqueous Micro Phase Separation: Construction of an Artificial Model of Cellular Assembly , 2019, Front. Chem..

[162]  T. Peter,et al.  Liquid-liquid phase separation and morphology of internally mixed dicarboxylic acids/ammonium sulfate/water particles , 2011 .

[163]  L. Benyahia,et al.  pH-responsive water-in-water Pickering emulsions. , 2015, Langmuir : the ACS journal of surfaces and colloids.

[164]  H. Stone,et al.  Flow-induced gelation of microfiber suspensions , 2017, Proceedings of the National Academy of Sciences.

[165]  R. Guo,et al.  Batch-Scale Preparation of Reverse Janus Emulsions. , 2019, Langmuir : the ACS journal of surfaces and colloids.

[166]  R. van Roij,et al.  Water-in-Water Emulsions Stabilized by Nanoplates. , 2015, ACS macro letters.

[167]  H. Shum,et al.  Suppressing the Folding of Flowing Viscous Jets Using an Electric Field , 2015 .

[168]  Dae Kun Hwang,et al.  Shrinking, growing, and bursting: microfluidic equilibrium control of water-in-water droplets. , 2016, Lab on a chip.

[169]  P. Fletcher,et al.  Water-in-water emulsions based on incompatible polymers and stabilized by triblock copolymers-templated polymersomes. , 2013, Langmuir : the ACS journal of surfaces and colloids.

[170]  L. Mei,et al.  Interfacial Tension of Poly(ethylene glycol) + Salt + Water Systems , 1996 .

[171]  Ho Cheung Shum,et al.  Forced generation of simple and double emulsions in all-aqueous systems , 2012, 1212.1084.

[172]  Alexander K. Buell,et al.  Protein microgels from amyloid fibril networks. , 2015, ACS nano.

[173]  B. Binks,et al.  Colloidal Particles at Liquid Interfaces: Particles at Curved Liquid Interfaces , 2006 .

[174]  B. V. Schmidt,et al.  Supramolecular Compartmentalized Hydrogels via Polydopamine Particle-Stabilized Water-in-Water Emulsions , 2019, Langmuir : the ACS journal of surfaces and colloids.

[175]  B. Mosadegh,et al.  Nanoliter Liquid Patterning in Aqueous Environments for Spatially-defined Reagent Delivery to Mammalian Cells Hhs Public Access , 2022 .

[176]  Armand Ajdari,et al.  Stability of a jet in confined pressure-driven biphasic flows at low reynolds numbers. , 2007, Physical review letters.

[177]  A. Bertram,et al.  Predicting the relative humidities of liquid-liquid phase separation, efflorescence, and deliquescence of mixed particles of ammonium sulfate, organic material, and water using the organic-to-sulfate mass ratio of the particle and the oxygen-to-carbon elemental ratio of the organic component , 2011 .

[178]  Gang Zhao,et al.  All-Aqueous-Phase Microfluidics for Cell Encapsulation. , 2019, ACS applied materials & interfaces.

[179]  Ho Cheung Shum,et al.  Osmo-solidification of all-aqueous emulsion with enhanced preservation of protein activity. , 2016, Journal of materials chemistry. B.

[180]  Shuichi Takayama,et al.  Aqueous biphasic microprinting approach to tissue engineering. , 2011, Biomicrofluidics.

[181]  RNA Granules Hitchhike on Lysosomes for Long-Distance Transport, Using Annexin A11 as a Molecular Tether , 2019, Cell.

[182]  P. Fernandes,et al.  Determination of partition coefficients of biomolecules in a microfluidic aqueous two phase system platform using fluorescence microscopy. , 2017, Journal of chromatography. A.

[183]  Selvaraj Raja,et al.  Aqueous Two Phase Systems for the Recovery of Biomolecules –A Review , 2012 .

[184]  F. Schönfeld,et al.  Protein diffusion across the interface in aqueous two-phase systems. , 2008, Langmuir : the ACS journal of surfaces and colloids.

[185]  B. Binks.  Colloidal particles at liquid interfaces. , 2008, Physical chemistry chemical physics : PCCP.

[186]  S. Tsai,et al.  Microfluidic magnetic self-assembly at liquid-liquid interfaces. , 2016, Soft matter.

[187]  Daeyeon Lee,et al.  AWE-somes: All Water Emulsion Bodies with Permeable Shells and Selective Compartments. , 2017, ACS applied materials & interfaces.

[188]  Sze Yi Mak,et al.  Diffusion-Dominated Pinch-Off of Ultralow Surface Tension Fluids. , 2019, Physical review letters.

[189]  Xiaobo Ji,et al.  Versatile preparation of nonspherical multiple hydrogel core PAM/PEG emulsions and hierarchical hydrogel microarchitectures. , 2014, Angewandte Chemie.

[190]  Wilhelm T S Huck,et al.  Fabrication of microgel particles with complex shape via selective polymerization of aqueous two-phase systems. , 2012, Small.

[191]  Jiaqu Chen,et al.  High-throughput-generating water-in-water droplet for monodisperse biocompatible particle synthesis , 2019, Journal of Materials Science.

[192]  A. Fernández-Nieves,et al.  Teaching Rayleigh–Plateau instabilities in the laboratory , 2015 .

[193]  A. Manz,et al.  Membrane-free electroextraction using an aqueous two-phase system , 2014 .

[194]  M. Kreutzer,et al.  Monodisperse hydrogel microspheres by forced droplet formation in aqueous two-phase systems. , 2011, Lab on a chip.

[195]  Gerald Muschiolik,et al.  Multiple emulsions for food use , 2007 .

[196]  Yuanjin Zhao,et al.  Emerging Droplet Microfluidics. , 2017, Chemical reviews.

[197]  S. Tomotika.  On the Instability of a Cylindrical Thread of a Viscous Liquid Surrounded by Another Viscous Fluid , 1935 .

[198]  V. N. Paunov,et al.  High throughput fabrication of cell spheroids by templating water-in-water Pickering emulsions , 2017 .

[199]  Philip C Bevilacqua,et al.  Bioreactor droplets from liposome-stabilized all-aqueous emulsions , 2014, Nature Communications.

[200]  Shuichi Takayama,et al.  Rounded multi-level microchannels with orifices made in one exposure enable aqueous two-phase system droplet microfluidics. , 2011, Lab on a chip.

[201]  M. Yamada,et al.  One-Step Formation of Microporous Hydrogel Sponges Encapsulating Living Cells by Utilizing Bicontinuous Dispersion of Aqueous Polymer Solutions. , 2019, ACS applied bio materials.

[202]  C. Monteux,et al.  Colloidal Particle Adsorption at Water-Water Interfaces with Ultralow Interfacial Tension. , 2017, Physical Review Letters.

[203]  T. Nicolai,et al.  Stabilization of Water-in-Water Emulsions by Linear Homo-Polyelectrolytes. , 2019, Langmuir : the ACS journal of surfaces and colloids.

[204]  Do Hyun Kim,et al.  Droplet-based microextraction in the aqueous two-phase system. , 2010, Journal of chromatography. A.

[205]  R. Lipowsky,et al.  Giant Vesicles Encapsulating Aqueous Two-Phase Systems: From Phase Diagrams to Membrane Shape Transformations , 2019, Front. Chem..

[206]  Shawn A. Chester,et al.  Printing ferromagnetic domains for untethered fast-transforming soft materials , 2018, Nature.

[207]  Paul D. Ashby,et al.  Compartmentalized, All-Aqueous Flow-Through-Coordinated Reaction Systems , 2019, Chem.

[208]  Taco Nicolai,et al.  Stabilization of water-in-water emulsions by addition of protein particles. , 2013, Langmuir : the ACS journal of surfaces and colloids.

[209]  Petra Schwille,et al.  Liposomes and polymersomes: a comparative review towards cell mimicking. , 2018, Chemical Society reviews.

[210]  R. Varadaraj,et al.  Colloid and interface science in the oil industry , 1996 .

[211]  Yu Wang,et al.  Colloids with valence and specific directional bonding , 2012, Nature.

[212]  S. Tsai,et al.  Microfluidic Generation of Particle-Stabilized Water-in-Water Emulsions. , 2017, Langmuir : the ACS journal of surfaces and colloids.

[213]  E. Kumacheva,et al.  Microfluidic encapsulation of cells in polymer microgels. , 2012, Small.

[214]  C. Keating,et al.  Microcompartmentation in artificial cells: pH-induced conformational changes alter protein localization. , 2010, Langmuir : the ACS journal of surfaces and colloids.

[215]  Kristin Robin Ko,et al.  Emerging Biotechnology Applications of Aqueous Two‐Phase Systems , 2018, Advanced healthcare materials.

[216]  Dino Di Carlo,et al.  Fabricating Shaped Microfibers with Inertial Microfluidics , 2014, Advanced materials.

[217]  Michiel T. Kreutzer,et al.  All-aqueous core-shell droplets produced in a microfluidic device , 2011 .

[218]  M. Nakano.  Places of emulsions in drug delivery. , 2000, Advanced drug delivery reviews.

[219]  I. M. Walker,et al.  The in vivo evaluation of emulsion formulations administered intramuscularly , 1987 .

[220]  Dino Di Carlo,et al.  Microtechnology for Cell Manipulation and Sorting. , 2017, Anticancer research.

[221]  C. Keating,et al.  Aqueous phase separation in giant vesicles. , 2002, Journal of the American Chemical Society.

[222]  I. Plazl,et al.  Experimental studies and modeling of α-amylase aqueous two-phase extraction within a microfluidic device , 2015 .

[223]  DoYeun Park,et al.  pH-Triggered Silk Fibroin/Alginate Structures Fabricated in Aqueous Two-Phase System. , 2019, ACS biomaterials science & engineering.

[224]  M. Přibyl,et al.  Electric field driven addressing of ATPS droplets in microfluidic chips , 2017 .

[225]  A. Hyman,et al.  Germline P Granules Are Liquid Droplets That Localize by Controlled Dissolution/Condensation , 2009, Science.

[226]  Ho Cheung Shum,et al.  On-Demand Droplet Collection for Capturing Single Cells. , 2020, Small.

[227]  H. Lekkerkerker,et al.  Decreased Interfacial Tension of Demixed Aqueous Polymer Solutions due to Charge. , 2015, Physical review letters.

[228]  P. Fernandes,et al.  Miniaturization of aqueous two‐phase extraction for biological applications: From micro‐tubes to microchannels , 2016, Biotechnology journal.

[229]  Ruben R G Soares,et al.  Partitioning in aqueous two‐phase systems: Analysis of strengths, weaknesses, opportunities and threats , 2015, Biotechnology journal.

[230]  L. Benyahia,et al.  Stabilization of Water-in-Water Emulsions by Polysaccharide-Coated Protein Particles. , 2016, Langmuir : the ACS journal of surfaces and colloids.

[231]  P. Bolhuis,et al.  Self-assembly of microcapsules via colloidal bond hybridization and anisotropy , 2015, Nature.

[232]  Stephen W. Michnick,et al.  Mechanisms and Consequences of Macromolecular Phase Separation , 2016, Cell.

[233]  Rajni Hatti-Kaul,et al.  Aqueous two-phase systems : methods and protocols , 2000 .

[234]  B. Murray,et al.  Whey protein microgel particles as stabilizers of waxy corn starch + locust bean gum water-in-water emulsions , 2016 .

[235]  Gorka Orive,et al.  Microcapsules and microcarriers for in situ cell delivery. , 2010, Advanced drug delivery reviews.

[236]  Yooli K Light,et al.  Rapid, continuous purification of proteins in a microfluidic device using genetically-engineered partition tags. , 2008, Lab on a chip.

[237]  João A P Coutinho,et al.  Aqueous biphasic systems: a boost brought about by using ionic liquids. , 2012, Chemical Society reviews.

[238]  Yuzuru Takamura,et al.  Cell separation by an aqueous two-phase system in a microfluidic device. , 2009, The Analyst.

[239]  T. Cubaud.  Wetting and lubricating film instabilities in microchannels , 2009 .

[240]  M. Antonietti,et al.  Water-in-Water Pickering Emulsion Stabilized by Polydopamine Particles and Cross-Linking. , 2018, Biomacromolecules.

[241]  Bernard P. Binks,et al.  Polymer-Protein Conjugate Particles with Biocatalytic Activity for Stabilization of Water-in-Water Emulsions. , 2017, ACS macro letters.

[242]  Shuichi Takayama,et al.  Aqueous Two Phase System Assisted Self-Assembled PLGA Microparticles , 2016, Scientific Reports.

[243]  C. Filipe,et al.  In vivo formation of protein based aqueous microcompartments. , 2009, Journal of the American Chemical Society.

[244]  D. Rousseau,et al.  Tailoring the morphology and rheology of phase-separated biopolymer gels using microbial cells as structure modifiers , 2014 .

[245]  David A Weitz,et al.  Drop formation in non-planar microfluidic devices. , 2012, Lab on a chip.

[246]  Frank A Fencl,et al.  Oil-Free Acoustofluidic Droplet Generation for Multicellular Tumor Spheroid Culture. , 2019, ACS applied bio materials.

[247]  M. Freedman.  Phase separation in organic aerosol. , 2017, Chemical Society reviews.

[248]  Aaron S. Meyer,et al.  Concentration of mammalian genomic DNA using two‐phase aqueous micellar systems , 2009, Biotechnology and bioengineering.

[249]  F. Hellman,et al.  Reconfigurable ferromagnetic liquid droplets , 2019, Science.

[250]  L. Benyahia,et al.  Influence of the Protein Particle Morphology and Partitioning on the Behavior of Particle-Stabilized Water-in-Water Emulsions. , 2016, Langmuir : the ACS journal of surfaces and colloids.

[251]  Ana M. Azevedo,et al.  Partitioning in Aqueous Two-Phase Systems: Fundamentals, Applications and Trends , 2016 .

[252]  Sze Yi Mak,et al.  Perturbation-induced droplets for manipulating droplet structure and configuration in microfluidics , 2015 .

[253]  B. Grzybowski,et al.  Systems of mechanized and reactive droplets powered by multi-responsive surfactants , 2018, Nature.

[254]  A. Abate,et al.  SiC-Seq: Single-cell genome sequencing at ultra high-throughput with microfluidic droplet barcoding , 2017, Nature Biotechnology.

[255]  S. Takayama,et al.  Aqueous Two‐Phase System Patterning of Microbubbles: Localized Induction of Apoptosis in Sonoporated Cells , 2013 .

[256]  Anthony A. Hyman,et al.  Biomolecular condensates: organizers of cellular biochemistry , 2017, Nature Reviews Molecular Cell Biology.

[257]  M. Zheng,et al.  DNA-controlled partition of carbon nanotubes in polymer aqueous two-phase systems. , 2014, Journal of the American Chemical Society.

[258]  Tiantian Kong,et al.  Spinning and Applications of Bioinspired Fiber Systems. , 2019, ACS nano.

[259]  M. J. Rosen.  Surfactants and Interfacial Phenomena , 1978 .

[260]  W. Hennink,et al.  The use of aqueous PEG/dextran phase separation for the preparation of dextran microspheres. , 1999, International journal of pharmaceutics.