Continuous, One-pot Synthesis and Post-Synthetic Modification of NanoMOFs Using Droplet Nanoreactors.

Metal-organic frameworks (MOFs); also known as porous coordination polymers (PCP) are a class of porous crystalline materials constructed by connecting metal clusters via organic linkers. The possibility of functionalization leads to virtually infinite MOF designs using generic modular methods. Functionalized MOFs can exhibit interesting physical and chemical properties including accelerated adsorption kinetics and catalysis. Although there are discrete methods to synthesize well-defined nanoscale MOFs, rapid and flexible methods are not available for continuous, one-pot synthesis and post-synthetic modification (functionalization) of MOFs. Here, we show a continuous, scalable nanodroplet-based microfluidic route that not only facilitates the synthesis of MOFs at a nanoscale, but also offers flexibility for direct functionalization with desired functional groups (e.g., -COCH3, fluorescein isothiocyanate; FITC). In addition, the presented route of continuous manufacturing of functionalized nanosized MOFs takes significantly less time compared to state-of-the-art batch methods currently available (1 hr vs. several days). We envisage our approach to be a breakthrough method for synthesizing complex functionalized nanomaterials (metal, metal oxides, quantum dots and MOFs) that are not accessible by direct batch processing and expand the range of a new class of functionalized MOF-based functional nanomaterials.

Evaluation of peristaltic micromixers for highly integrated microfluidic systems.

Microfluidic devices based on the multilayer soft lithography allow accurate manipulation of liquids, handling reagents at the sub-nanoliter level, and performing multiple reactions in parallel processors by adapting micromixers. Here, we have experimentally evaluated and compared several designs of micromixers and operating conditions to find design guidelines for the micromixers. We tested circular, triangular, and rectangular mixing loops and measured mixing performance according to the position and the width of the valves that drive nanoliters of fluids in the micrometer scale mixing loop. We found that the rectangular mixer is best for the applications of highly integrated microfluidic platforms in terms of the mixing performance and the space utilization. This study provides an improved understanding of the flow behaviors inside micromixers and design guidelines for micromixers that are critical to build higher order fluidic systems for the complicated parallel bio/chemical processes on a chip.

Microfluidics for Antibiotic Susceptibility and Toxicity Testing.

The recent emergence of antimicrobial resistance has become a major concern for worldwide policy makers as very few new antibiotics have been developed in the last twenty-five years. To prevent the death of millions of people worldwide, there is an urgent need for a cheap, fast and accurate set of tools and techniques that can help to discover and develop new antimicrobial drugs. In the past decade, microfluidic platforms have emerged as potential systems for conducting pharmacological studies. Recent studies have demonstrated that microfluidic platforms can perform rapid antibiotic susceptibility tests to evaluate antimicrobial drugs' efficacy. In addition, the development of cell-on-a-chip and organ-on-a-chip platforms have enabled the early drug testing, providing more accurate insights into conventional cell cultures on the drug pharmacokinetics and toxicity, at the early and cheaper stage of drug development, i.e., prior to animal and human testing. In this review, we focus on the recent developments of microfluidic platforms for rapid antibiotics susceptibility testing, investigating bacterial persistence and non-growing but metabolically active (NGMA) bacteria, evaluating antibiotic effectiveness on biofilms and combinatorial effect of antibiotics, as well as microfluidic platforms that can be used for in vitro antibiotic toxicity testing.

Cell-based dose responses from open-well microchambers.

Cell-based assays play a critical role in discovery of new drugs and facilitating research in cancer, immunology, and stem cells. Conventionally, they are performed in Petri dishes, tubes, or well plates, using milliliters of reagents and thousands of cells to obtain one data point. Here, we are introducing a new platform to realize cell-based assay capable of increased throughput and greater sensitivity with a limited number of cells. We integrated an array of open-well microchambers into a gradient generation system. Consequently, cell-based dose responses were examined with a single device. We measured IC50 values of three cytotoxic chemicals, Triton X-100, H2O2, and cadmium chloride, as model compounds. The present system is highly suitable for the discovery of new drugs and studying the effect of chemicals on cell viability or mortality with limited samples and cells.

Log-scale dose response of inhibitors on a chip.

We demonstrate the accommodation of log-scale concentration gradients of inhibitors on a single microfluidic chip with a semidirect dilution capability of reagents for the determination of the half-inhibitory concentration or IC(50). The chip provides a unique tool for hosting a wide-range of concentration gradient for studies that require an equal distribution of measuring points on a logarithmic scale. Using Matrix metalloproteinase IX and three of its inhibitors, marimastat, batimastat, and CP471474, we evaluated the IC(50) of each inhibitor with a single experiment. The present work could be applied to the systematic study of biochemical binding and inhibition processes particularly in the field of mechanistic enzymology and the pharmaceutical industry.

Creation of stepwise concentration gradient in picoliter droplets for parallel reactions of matrix metalloproteinase II and IX.

We present a new methodology for generating a stepwise concentration gradient in a series of microdroplets by using monolithic micro valves that act as "faucets" in micrometer-scale. A distinct concentration gradient of a substrate was generated for the determination of the kinetic parameters of two different enzymes using only 10 picoliter-scale droplets. With a single experiment on a chip, we obtained K(M) and k(cat) values of matrix metalloproteinase 2 (MMP-2) and matrix metalloproteinase 9 (MMP-9), and compared the catalytic competence of the two enzymes. The present system and method are highly suitable for applications where the reagents or samples are limited and precious.

Generating nonlinear concentration gradients in microfluidic devices for cell studies.

We describe a microfluidic device for generating nonlinear (exponential and sigmoidal) concentration gradients, coupled with a microwell array for cell storage and analysis. The device has two inputs for coflowing multiple aqueous solutions, a main coflow channel and an asymmetrical grid of fluidic channels that allows the two solutions to combine at intersection points without fully mixing. Due to this asymmetry and diffusion of the two species in the coflow channel, varying amounts of the two solutions enter each fluidic path. This induces exponential and sigmoidal concentration gradients at low and high flow rates, respectively, making the microfluidic device versatile. A key feature of this design is that it is space-saving, as it does not require multiplexing or a separate array of mixing channels. Furthermore, the gradient structure can be utilized in concert with cell experiments, to expose cells captured in microwells to various concentrations of soluble factors. We demonstrate the utility of this design to assess the viability of fibroblast cells in response to a range of hydrogen peroxide (H(2)O(2)) concentrations.

Determination of kinetic parameters, Km and kcat, with a single experiment on a chip.

We have demonstrated a multistep enzyme reaction on a chip to determine the key kinetic parameters of enzyme reaction. We designed and fabricated a fully integrated microfluidic chip to have sample metering, mixing, and incubation functionalities. The chip generates a gradient of reagent concentrations in 11 parallel processors. We used beta-galactosidase and its substrate, resorufin-beta-D-galactopyranoside, as the model system of the enzyme reaction. With a single experiment on the chip, we determined the key parameters for the enzyme kinetics, K(m) and k(cat), and evaluated the effect of inhibitor concentrations on the reaction rates. This study provides a new tool for evaluating various effectors, such as inhibitors and cofactors, on the initial rate of an enzyme reaction, and it could be applied to a comprehensive bio/chemical reaction study.