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Two covalent organic frameworks synthesized from flexible building blocks displayed good porosity, crystallinity, and high density of N and O atoms in the skeleton, which explained the high iodine capture ability at 5.51 g g−1.

Two pyridine-based conjugated microporous polymers with high micro-porosity exhibited a high CO2 capture value via weak supramolecular interaction.

Robust and emissive COFs via intramolecular hydrogen bond interaction suggested high sensitivity, selectivity, and sensibility towards 2,4,6-trinitrophenol.

Chemotaxis is a fundamental bacterial response mechanism to changes in chemical gradients of specific molecules known as chemoattractant or chemorepellent. The advancement of biological platforms for bacterial chemotaxis research is of significant interest for a wide range of biological and environmental studies. Many microfluidic devices have been developed for its study, but challenges still remain that can obscure analysis. For example, cell migration can be compromised by flow-induced shear stress, and bacterial motility can be impaired by nonspecific cell adhesion to microchannels. Also, devices can be complicated, expensive, and hard to assemble. We address these issues with a three-channel microfluidic platform integrated with natural biopolymer membranes that are assembled in situ. This provides several unique attributes. First, a static, steady and robust chemoattractant gradient was generated and maintained. Second, because the assembly incorporates assembly pillars, the assembled membrane arrays connecting nearby pillars can be created longer than the viewing window, enabling a wide 2D area for study. Third, the in situ assembled biopolymer membranes minimize pressure and/or chemiosmotic gradients that could induce flow and obscure chemotaxis study. Finally, nonspecific cell adhesion is avoided by priming the polydimethylsiloxane (PDMS) microchannel surfaces with Pluronic F-127. We demonstrated chemotactic migration of Escherichia coli as well as Pseudomonas aeruginosa under well-controlled easy-to-assemble glucose gradients. We characterized motility using the chemotaxis partition coefficient (CPC) and chemotaxis migration coefficient (CMC) and found our results consistent with other reports. Further, random walk trajectories of individual cells in simple bright field images were conveniently tracked and presented in rose plots. Velocities were calculated, again in agreement with previous literature. We believe the biopolymer membrane-integrated platform represents a facile and convenient system for robust quantitative assessment of cellular motility in response to various chemical cues.

The integration of membranes in microfluidic devices has been extensively exploited for various chemical engineering and bioengineering applications over the past few decades. To augment the applicability of membrane-integrated microfluidic platforms for biomedical and tissue engineering studies, a biologically friendly fabrication process with naturally occurring materials is highly desired. The in situ preparation of membranes involving interfacial reactions between parallel laminar flows in microfluidic networks, known as the flow-assembly technique, is one of the most biocompatible approaches. Membranes of many types with flexible geometries have been successfully assembled inside complex microchannels using this facile and versatile flow-assembly approach. Chitosan is a naturally abundant polysaccharide known for its pronounced biocompatibility, biodegradability, good mechanical stability, ease of modification and processing, and film-forming ability under near-physiological conditions. Chitosan membranes assembled by flows in microfluidics are freestanding, robust, semipermeable, and well-aligned in microstructure, and show high affinity to bioactive reagents and biological components (e.g. biomolecules, nanoparticles, or cells) that provide facile biological functionalization of microdevices. Here, we discuss the recent developments and optimizations in the flow-assembly of chitosan membranes and chitosan-based membranes in microfluidics. Furthermore, we recapitulate the applications of the chitosan membrane-integrated microfluidic platforms dedicated to biology, biochemistry, and drug release fields, and envision the future developments of this important platform with versatile functions.

Flow-assembled chitosan membranes are robust and semipermeable hydrogel structures formed in microfluidic devices that have been used for important applications such as gradient generation and studying cell-cell signaling. One challenge, however, remains unresolved. When a polydimethylsiloxane (PDMS) microchannel with a flow-assembled, deprotonated chitosan membrane (DCM) is treated with anti-adhesion agents such as Pluronic F-127 to prevent biomolecular and cellular adsorption on PDMS, the interaction between DCM and PDMS is compromised and the DCM easily delaminates. To address this challenge, DCMs in microfluidics are crosslinked with glutaraldehyde to modulate their properties, and the altered properties of the glutaraldehyde treated chitosan membrane (GTCM) are investigated. First, the GTCM's acidic resistance was confirmed, its mechanical robustness against hydrostatic pressure was significantly improved, and it remained intact on PDMS after Pluronic treatment. Second, crystallization in DCM and GTCM was investigated with quantitative polarized light microscopy (qPLM), which revealed that GTCM's optical retardance and anisotropy were lower, implying less molecular alignment than in DCM. Finally, membrane permeability was tested with FITC-labeled dextran transport experiments, which showed that the transport across GTCM was slightly higher than that across DCM. Overall, glutaraldehyde-crosslinked chitosan membrane has better acidic resistance, higher strength under Pluronic treatment, and less molecular microalignment, while its semi-permeability is retained. This study demonstrates how glutaraldehyde crosslinking can be used to modify and improve biopolymer membrane properties for broader applications, such as in an acidic environment or when Pluronic passivation is needed.

Biopolymer membranes assembled in microfluidic devices offer many biological process- and analysis-related applications. One of the key characteristics of bio-fabricated membranes is their porosity, which regulates the transport of molecules, ions, or particles and contributes to their semi-permeability and selectivity. This study aims to tune the porosity of biofabricated chitosan membranes (CM) using incorporated nanoparticles as templates. CM with polystyrene nanoparticles (CM-np) were assembled by flow in microchannel networks. The membranes with incorporated nanoparticles were crosslinked with glutaraldehyde, and then the nanoparticles were dissolved with dimethyl sulfoxide. The in situ synthesized porous CM (pCM) were characterized with scanning electron microscopy and polarized light microscopy. Permeability tests confirmed the increased pore sizes of the pCM and enhanced permeability to macromolecules. Sharper static gradients in three-channel microfluidic devices were demonstrated with the pCM as compared to those with the original CM. The capability to customize the porosity of flow-assembled, freestanding and robust biopolymer membranes inside a microfluidic network is attractive and broadens the applications of these membranes in biomolecular and cellular studies.

Membrane functionality is crucial in microfluidics for realizing operations such as filtration, separation, concentration, signaling among cells and gradient generation. Currently, common methods often sandwich commercially available membranes in multi-layer devices, or use photopolymerization or temperature-induced gelation to fabricate membrane structures in one-layer devices. Biofabrication offers an alternative to forming membrane structures with biomimetic materials and mechanisms in mild conditions. We have recently developed a biofabrication strategy to form parallel biopolymer membranes in gas-permeable polydimethylsiloxane (PDMS) microfluidic devices, which used positive pressure to dissipate air bubbles through PDMS to initiate membrane formation but required careful pressure balancing between two flows. Here, we report a technical innovation by simply placing as needed an add-on PDMS vacuum layer on PDMS microfluidic devices to dissipate air bubbles and guide the biofabrication of biopolymer membranes. Vacuuming through PDMS was simply achieved by either withdrawing a syringe or releasing a squeezed nasal aspirator. Upon vacuuming, air bubbles dissipated within minutes, membranes were effortlessly formed, and the add-on vacuum layer can be removed. Subsequent membrane growth could be robustly controlled with the flows and pH of solutions. This new process is user-friendly and has achieved a 100% success rate in more than 200 trials in membrane biofabrication.

We report an in situ biofabrication strategy that conveniently partitions microfluidic networks into physically separated while chemically communicating microchannels with semipermeable biopolymer membranes, which enable the facile generation of static gradients for biomedical applications. The biofabrication of parallel biopolymer membranes was initiated with the dissipation of trapped air bubbles in parallel apertures in polydimethylsiloxane (PDMS) microfluidic devices, followed by tunable membrane growth with precise temporal and spatial control to the desired thickness. Static gradients were generated within minutes and well maintained over time by pure diffusion of molecules through the biofabricated semipermeable membranes. As an example application, the static gradient of alpha factor was generated to study the development of the "shmoo" morphology of yeast over time. The in situ biofabrication provides a simple approach to generate static gradients and an ideal platform for biological applications where flow-free static gradients are indispensable.

This article describes the synthesis and carbon dioxide uptake of new organic microporous frameworks with built-in metal sites in the skeleton. Three novel microporous polymers have been synthesized via a Sonogashira–Hagihara coupling reaction with 1,3,5-triethynylbenzene and diverse metallosalen building blocks. These materials are insoluble in conventional solvents and exhibit high thermal and chemical stability. According to the nitrogen physisorption isotherms, the highest Brunauer–Emmett–Teller specific surface area up to 1200 m2 g−1 was obtained for three polymer frameworks with a pore volume of 0.94 cm3 g−1. The polymer frameworks display high carbon dioxide uptake capacities (up to 8.2 wt%) and good selectivity at 273 K and 1 bar, which is impacted on significantly by the porosity of the frameworks, active heteroatoms and coordinatively unsaturated metal sites in the skeletons.

A multifunctional carbazole-based conjugated microporous polymer MFCMP-1 is successfully prepared by oxidative coupling polymerization using a single monomer and structurally characterized. A new three-dimensional π-conjugated polymer framework can be combined with permanent microporous, highly luminescent properties and abundant nitrogen activated sites in the skeleton. It possesses a large BET surface area of over 840 m2 g−1 with a pore volume of 0.52 cm3 g−1, and displays a high carbon dioxide uptake capacity (up to 3.69 mmol g−1) at 273 K and 1 bar, with good selectivity towards CO2 over N2 and CH4. MFCMP-1 exhibits also strong fluorescent emission at 529 nm after excitation at 380 nm in THF solution and works as a luminescent chemosensor towards hazardous and explosive molecules, such as nitrobenzene, 2-nitrotoluene, and 2,4-dinitrotoluene. In addition, MFCMP-1 features a high concentration of Lewis base nitrogen sites on its internal surfaces; it thus acts as a highly efficient recyclable heterogeneous organocatalyst towards Knoevenagel reaction of malononitrile with aromatics, heterocyclic aldehydes, and cyclic ketones. Furthermore, we further highlight that the ease of synthesis and low cost, coupled with multifunctional properties, make MFCMP-1 an attractive functional material in practical applications.

A new porphyrin-based conjugated microporous polymer with rich nitrogen sites in the skeleton has been synthesized by alkyne–alkyne homocoupling reaction. The Brunauer–Emmett–Teller specific surface area up to 662 m2 g−1 was obtained for the new polymer framework with a pore volume of 0.55 cm3 g−1. The polymer network displays high carbon dioxide uptake capacity (up to 3.58 mmol g−1) at 273 K and 1 bar, with good selectivity towards CO2 over N2 and CH4. Furthermore, this framework also acts as a solid organocatalyst towards Knoevenagel reaction of malononitrile with aromatic, heterocyclic aldehydes, and cyclic ketones. The reaction afforded the corresponding products in excellent yields (up to 99%) with short times. Moreover, the heterogeneous catalyst was also found to exhibit an excellent recyclability (up to 10 times) without loss of efficiency.

Because of their stimuli-responsiveness to chemical and pH gradients, polysaccharide hydrogels such as chitosan and alginate can be assembled as scaffolds for biomolecules or cells. Using the electrical and flow control available in microfluidic networks, in situ fabrication of 3D hydrogel scaffolds can be programmed in space and time to arrange biological components as an in vitro biochemically communicating system. Flexible in situ on-demand construction of a biocompatible scaffold within microfluidics holds promise for the assembly of biological components and systems for in vitro analysis and investigation. We foresee a wide spectrum applications ranging from replication of metabolic pathways as testbeds for drug discovery to identification of cell signaling mechanisms and observation of cellular response.

Programmable 3D cell assembly under physiological pH conditions is achieved using electrodeposited stimuli-responsive alginate gels in a microfluidic device, with parallel sidewall electrodes enabling direct observation of the cell assembly. Electrically triggered assembly and subsequent viability of mammalian cells is demonstrated, along with spatially programmable, multi-address assembly of different strains of E. coli cells. Our approach enables in vitro study of dynamic cellular and inter-cellular processes, from cell growth and stimulus/response to inter-colony and inter-species signaling.

Stimuli-responsive polysaccharides that can undergo a sol–gel transition in response to localized electrical signals provide a unique opportunity to electroaddress biological components at device interfaces. Most polysaccharide electroaddressing mechanisms use electrochemical reactions to generate pH gradients that can locally neutralize the polysaccharide and induce its reversible sol–gel transition to form a hydrogel film adjacent to the electrode surface. The calcium-responsive polysaccharide alginate is an exception; it may electrodeposit without requiring extreme pH gradients and thus may provide a means to electroaddress pH-sensitive biological components. Here, we use a novel device to characterize the mechanism for the anodic electrodeposition of a calcium alginate hydrogel. This device consists of a transparent fluidic channel with built-in sidewall electrodes that allows Ca–alginate electrodeposition to be directly measured by non-destructive optical and spectroscopic methods. We hypothesize a 3-step mechanism for calcium–alginate electrodeposition: (i) water is electrolyzed to locally generate protons (or hydronium ions); (ii) these protons are consumed by reacting with suspended CaCO3 particles and this “buffering” reaction generates a gradient in soluble Ca2+; and (iii) the locally generated Ca2+ ions interact with alginate to induce its sol–gel transition. We verified this electrodeposition mechanism using pH-responsive dyes to observe the local pH gradients during gel formation, Ca2+ indicator dyes to observe the Ca2+ gradient, and in situ Raman spectroscopy to demonstrate a strong interaction between soluble Ca2+ and alginate. Importantly, these results demonstrate electrodeposition without the need for a substantial pH excursion from neutrality. Thus, calcium alginate appears especially well-suited for electroaddressing labile biological components for applications in biosensors, biofabrication and BioMEMS.

The emergence of bacteria that evade antibiotics has accelerated research on alternative approaches that do not target cell viability. One such approach targets cell-cell communication networks mediated by small molecule signaling. In this report, we assemble biological nanofactories within a bioMEMS device to capture and manipulate the behavior of quorum sensing (QS) bacteria as a step toward modifying small molecule signaling. Biological nanofactories are bio-inspired nanoscale constructs which can include modules with different functionalities, such as cell targeting, molecular sensing, product synthesis, and ultimately self-destruction. The biological nanofactories reported here consist of targeting, sensing, synthesis and, importantly, assembly modules. A bacteria-specific antibody constitutes the targeting module while a genetically engineered fusion protein contains the sensing, synthesis and assembly modules. The nanofactories are assembled on chitosan electrodeposited within a microchannel of the bioMEMS device; they capture QS bacteria in a spatially selective manner and locally synthesize and deliver the "universal" small signaling molecule autoinducer-2 (AI-2) at the captured cell surface. The nanofactory based AI-2 delivery is demonstrated to alter the progression of the native AI-2 based QS response of the captured bacteria. Prospects are envisioned for utilizing our technique as a test-bed for understanding the AI-2 based QS response of bacteria as a means for developing the next generation of antimicrobials.

We report the first in situ quantitative visualization and characterization of electro-induced chitosan hydrogel growth in an aqueous environment. This was enabled with a pair of sidewall electrodes within a transparent fluidic system, which allowed us to resolve the electrogelling mechanism and interpret the dominant causes responsible for the formation and density distribution of the deposited hydrogel. The pH and the time-dependent growth profiles of the chitosan hydrogel were directly visualized, analyzed, and characterized. The results indicate that the gelation and immobilization of chitosan onto the cathode at a pH above its pKa value (∼6.3) are due to the electrochemically generated concentration gradient of reactant OH− ions, and their subsequent neutralization of the NH3+ groups of chitosan chains in solution near the cathode. The increased gel density around the fringes of the electrodes was demonstrated and correlated with the electrophoretic migration of chitosan cations during deposition. Simulation of the electric potential/field distribution, together with the corresponding dry film topography confirmed the non-uniform, electric field-dependent density distribution of deposited hydrogel. This report provides fundamental understanding towards the mechanism and the kinetics of the electro-induced chitosan gel formation. It also provides important guidelines for pursuing its application in bio-components integrated microsystems. The method in use exemplifies a simple, effective and non-destructive approach for in situ characterization of electro-responsive biopolymers in an aqueous environment.

Chitosan is a naturally derived polymer with applications in a variety of industrial and biomedical fields. Recently, it has emerged as a promising material for biological functionalization of microelectromechanical systems (bioMEMS). Due to its unique chemical properties and film forming ability, chitosan serves as a matrix for the assembly of biomolecules, cells, nanoparticles, and other substances. The addition of these components to bioMEMS devices enables them to perform functions such as specific biorecognition, enzymatic catalysis, and controlled drug release. The chitosan film can be integrated in the device by several methods compatible with standard microfabrication technology, including solution casting, spin casting, electrodeposition, and nanoimprinting. This article surveys the usage of chitosan in bioMEMS to date. We discuss the common methods for fabrication, modification, and characterization of chitosan films, and we review a number of demonstrated chitosan-based microdevices. We also highlight the advantages of chitosan over some other functionalization materials for micro-scale devices.

We report a biofunctionalization strategy for the assembly of catalytically active enzymes within a completely packaged bioMEMS device, through the programmed generation of electrical signals at spatially and temporally defined sites. The enzyme of a bacterial metabolic pathway, S-adenosylhomocysteine nucleosidase (Pfs), is genetically fused with a pentatyrosine "pro-tag" at its C-terminus. Signal responsive assembly is based on covalent conjugation of Pfs to the aminopolysaccharide, chitosan, upon biochemical activation of the pro-tag, followed by electrodeposition of the enzyme-chitosan conjugate onto readily addressable sites in microfluidic channels. Compared to traditional physical entrapment and surface immobilization approaches in microfluidic environments, our signal-guided electrochemical assembly is unique in that the enzymes are assembled under mild aqueous conditions with spatial and temporal programmability and orientational control. Significantly, the chitosan-mediated enzyme assembly can be reversed, making the bioMEMS reusable for repeated assembly and catalytic activity. Additionally, the assembled enzymes retain catalytic activity over multiple days, demonstrating enhanced enzyme stability. We envision that this assembly strategy can be applied to rebuild metabolic pathways in microfluidic environments for antimicrobial drug discovery.

We report facile in situ biomolecule assembly at readily addressable sites in microfluidic channels after complete fabrication and packaging of the microfluidic device. Aminopolysaccharide chitosan's pH responsive and chemically reactive properties allow electric signal-guided biomolecule assembly onto conductive inorganic surfaces from the aqueous environment, preserving the activity of the biomolecules. A transparent and nonpermanently packaged device allows consistently leak-free sealing, simple in situ and ex situ examination of the assembly procedures, fluidic input/outputs for transport of aqueous solutions, and electrical ports to guide the assembly onto the patterned gold electrode sites within the channel. Both in situ fluorescence and ex situ profilometer results confirm chitosan-mediated in situ biomolecule assembly, demonstrating a simple approach to direct the assembly of biological components into a completely fabricated device. We believe that this strategy holds significant potential as a simple and generic biomolecule assembly approach for future applications in complex biomolecular or biosensing analyses as well as in sophisticated microfluidic networks as anticipated for future lab-on-a-chip devices.