Equal contribution

The controlled self-assembly of polymer-stabilized quantum dots (QDs) into mesoscale aqueous spherical assemblies termed quantum dot compound micelles (QDCMs) using a two-phase gas-segmented microfluidic reactor is described. Self-assembly is initiated by the fast mixing of water (approximately 1 s) with a blend solution of polystyrene-coated QDs and amphiphilic polystyrene-block-poly(acrylic acid) stabilizing chains via chaotic advection within liquid plugs moving through a sinusoidal channel. Subsequent recirculating flow within a post-formation channel subjects the dynamic QDCMs to shear-induced processing, controlled via the flow rate and channel length, before a final quench into pure water. During processing, larger QDCMs within the initial population undergo breakup into smaller particles, resulting in smaller mean particle sizes, smaller relative standard deviations, and more skewed distribution shapes, as the overall shear exposure is increased. For these cases, shear-induced size reduction is sufficient to dominate surface tension-driven growth.

The controlled self-assembly of large compound quantum dot micelles (QDCMs), consisting of constituents of polymer-stabilized quantum dots (QDs) and amphiphilic polystyrene-b-poly(acrylic acid) stabilizing chains, in gas-liquid segmented microfluidic reactors is demonstrated. Self-assembly is initiated by fast mixing of water with the polymer constituents via chaotic advection, as liquid plugs containing reactants move through a sinusoidal mixing channel. The resulting QDCMs are then processed within a postformation channel, where circulating flow patterns develop within the liquid plugs, followed by off-chip quenching and analysis by transmission electron microscopy (TEM). Particle processing via circulating flow is found to involve a combination of particle growth via collision-induced coalescence and shear-induced particle breakup. The final mean QDCM sizes represent kinetic states arising from the competition between these two mechanisms, depending on tunable chemical and flow parameters. A systematic investigation of the experimental variables that influence particle size and polydispersity, including water concentration, flow rate, and the gas-to-liquid flow ratio, is conducted, demonstrating tunability of QDCM sizes in the range of approximately 40-140 nm. The importance of shear-induced particle breakup in the limit of high shear is illustrated by a common minimum particle size, 41 +/- 1 nm, which is achieved for all water contents by increasing the total flow rate to sufficiently high values.

Hydrodynamic phenomena are ubiquitous in living organisms and can be used to manipulate cells or emulate physiological microenvironments experienced in vivo. Hydrodynamic effects influence multiple cellular properties and processes, including cell morphology, intracellular processes, cell–cell signaling cascades and reaction kinetics, and play an important role at the single-cell, multicellular, and organ level. Selected hydrodynamic effects can also be leveraged to control mechanical stresses, analyte transport, as well as local temperature within cellular microenvironments. With a better understanding of fluid mechanics at the micrometer-length scale and the advent of microfluidic technologies, a new generation of experimental tools that provide control over cellular microenvironments and emulate physiological conditions with exquisite accuracy is now emerging. Accordingly, we believe that it is timely to assess the concepts underlying hydrodynamic control of cellular microenvironments and their applications and provide some perspective on the future of such tools in in vitro cell-culture models. Generally, we describe the interplay between living cells, hydrodynamic stressors, and fluid flow-induced effects imposed on the cells. This interplay results in a broad range of chemical, biological, and physical phenomena in and around cells. More specifically, we describe and formulate the underlying physics of hydrodynamic phenomena affecting both adhered and suspended cells. Moreover, we provide an overview of representative studies that leverage hydrodynamic effects in the context of single-cell studies within microfluidic systems.

Local (cell-level) signaling environments, regulated by autocrine and paracrine signaling, and modulated by cell organization, are hypothesized to be fundamental stem cell fate control mechanisms used during development. It has, however, been challenging to demonstrate the impact of cell-level organization on stem cell fate control and to relate stem cell fate outcomes to autocrine and paracrine signaling. We address this fundamental problem using a combined in silico and experimental approach in which we directly manipulate, using laminar fluid flow, the local impact of endogenously secreted gp130-activating ligands and their activation of signal transducer and activator of transcription3 (STAT3) signaling in mouse embryonic stem cells (mESC). Our model analysis predicted that flow-dependent changes in autocrine and paracrine ligand binding would impact heterogeneity in cell-and colony-level STAT3 signaling activation and cause a gradient of cell fate determination along the direction of flow. Interestingly, analysis also predicted that local cell density would be inversely proportional to the degree to which endogenous secretion contributed to cell fate determination. Experimental validation using functional activation of STAT3 by secreted factors under microfluidic perfusion culture demonstrated that STAT3 activation and consequently mESC fate were manipulable by flow rate, position in the flow field, and local cell organization. As a unique demonstration of how quantitative control of autocrine and paracrine signaling can be integrated with spatial organization to elicit higher order cell fate effects, this work provides a general template to investigate organizing principles due to secreted factors.Brownian dynamics simulation | embryonic stem cells | leukemia inhibitory factor | cell fate control | cell heterogeneity T he developing embryo uses many spatially and temporally regulated mechanisms of signal propagation, including autocrine, paracrine, and extracellular matrix-mediated signals to control the proliferation and differentiation of progenitor cells. Based on our understanding of in vivo development, in vitro strategies attempt to mimic developmental mechanisms and establish artificial environments or niches that promote specific cell fates. Despite considerable progress in our ability to control pluripotent cell fate in vitro, significant variability and heterogeneity in differentiation protocols exist. One reason for this is thought to be our inability to mimic the temporally and spatially diverse signaling environments that occur in the embryo. Herein we use mouse embryonic stem cells (mESC) cultures as a model system to reveal a role for autocrine and paracrine signaling in cell fate control and to demonstrate how endogenous signaling and cell organization interact to create higher order local cellular environments, or niches.Mouse ESC cultures normally include the soluble interleukin-6 (IL-6) ligand family member leukemia inhibitory factor (LIF). IL-6 ligands such as LIF signal through the Jan...