High resolution fabrication of nanostructures using controlled proximity nanostencil lithography

Nanostencil lithography has a number of distinct benefits that make it an attractive nanofabrication processes, but the inability to fabricate features with nanometer precision has significantly limited its utility. In this paper, we describe a nanostencil lithography process that provides sub-15 nm resolution even for 40-nm thick structures by using a sacrificial layer to control the proximity between the stencil and substrate, thereby enhancing the correspondence between nanostencil patterns and fabricated nanostructures. We anticipate that controlled proximity nanostencil lithography will provide an environmentally stable, clean, and positive-tone candidate for fabrication of nanostructures with high resolution.United States. Air Force (Contract FA8721-05-C-0002

Enhanced discrimination of DNA molecules in nanofluidic channels through multiple measurements

Author Manuscript 2013 March 21.Nanofluidic sensing elements have been the focus of recent experiments for numerous applications ranging from nucleic acid fragment sizing to single-molecule DNA sequencing. These applications critically rely on high measurement fidelity, and methods to increase resolution are required. Herein, we describe fabrication and testing of a nanochannel device that enhances measurement resolution by performing multiple measurements (>100) on single DNA molecules. The enhanced measurement resolution enabled length discrimination between a mixture of λ-DNA (48.5 kbp) and T7 DNA (39.9 kbp) molecules, which were detected as transient current changes during translocation of the molecules through the nanochannel. As long DNA molecules are difficult to resolve quickly and with high fidelity with conventional electrophoresis, this approach may yield potentially portable, direct electrical sizing of DNA fragments with high sensitivity and resolution.National Institutes of Health (U.S.) (Grant R21EB009180)United States. Air Force (Contract FA8721-05-C- 0002

Integration of Solid-State Nanopores in Microfluidic Networks via Transfer Printing of Suspended Membranes

Solid-state nanopores have emerged as versatile single-molecule sensors for applications including DNA sequencing, protein unfolding, micro-RNA detection, label-free detection of single nucleotide polymorphisms, and mapping of DNA-binding proteins involved in homologous recombination. While machining nanopores in dielectric membranes provides nanometer-scale precision, the rigid silicon support for the membrane contributes capacitive noise and limits integration with microfluidic networks for sample preprocessing. Herein, we demonstrate a technique to directly transfer solid-state nanopores machined in dielectric membranes from a silicon support into a microfluidic network. The resulting microfluidic-addressable nanopores can sense single DNA molecules at high bandwidths and with low noise, owing to significant reductions in membrane capacitance. This strategy will enable large-scale integration of solid-state nanopores with microfluidic upstream and downstream processing and permit new functions with nanopores such as complex manipulations for multidimensional analysis and parallel sensing in two and three-dimensional architectures.National Institutes of Health (U.S.) (Grant R21EB009180)United States. Air Force (Contract FA8721-05-C-0002

A comprehensive review on biofuels from oil palm empty bunch (Efb): current status, potential, barriers and way forward

Biomass is an important renewable energy resource which primarily contributes to heating and cooling end use sectors. It is also a promising alternative source of biofuels to replace the depleting supply of fossil fuels. Surprisingly, few writers have been able to draw on the feedstock significance for oil palm empty fruit bunch (EFB) as the biomass resource for biofuels compared to the other types of biomass waste. Therefore, this paper presents a comprehensive review of EFB as a biomass resource presented in four major parts. First, the introduction covers the demand for bio-oil and describes the different kinds of feedstock, the relevance and potential of EFB biomass. Sec-ond, the characteristics of biomass are explained before it is upgraded as biofuel, drawing similarities and contrasts between EFB and other sources of biomass. Pyrolysis processes and reactors used for EFB conversion are described, and the factors affecting the bio-oil yield and quality are dis-cussed. Major reactor parameters are summarized and reactor optimization is discussed. Third, comparison on the properties of the bio-oil vs. petroleum in transportation, power generation, and heating are compared followed by prioritizing the bio-oil properties from the most to least critical, revealing the most promising methods for upgrading. Fourth, the environmental impact, including CO2 emission, of the use of EFB as a promising renewable energy resource and a cleaner alternative fuel is recommended. This paper has comprehensively reviewed the conversion of oil palm empty fruit bunches into biofuels, including the similarities and differences between biomasses, the best reactors, its comparison with fossil fuels, and bio-oil upgrading methods. The upgrading mapping matrix is created to present the best upgrading strategies for the optimum quality of biofuels. This paper serves as a one-stop center for EFB conversion into biofuels

Microfluidic technologies for accelerating the clinical translation of nanoparticles

Using nanoparticles for therapy and imaging holds tremendous promise for the treatment of major diseases such as cancer. However, their translation into the clinic has been slow because it remains difficult to produce nanoparticles that are consistent 'batch-to-batch', and in sufficient quantities for clinical research. Moreover, platforms for rapid screening of nanoparticles are still lacking. Recent microfluidic technologies can tackle some of these issues, and offer a way to accelerate the clinical translation of nanoparticles. In this Progress Article, we highlight the advances in microfluidic systems that can synthesize libraries of nanoparticles in a well-controlled, reproducible and high-throughput manner. We also discuss the use of microfluidics for rapidly evaluating nanoparticles in vitro under microenvironments that mimic the in vivo conditions. Furthermore, we highlight some systems that can manipulate small organisms, which could be used for evaluating the in vivo toxicity of nanoparticles or for drug screening. We conclude with a critical assessment of the near- and long-term impact of microfluidics in the field of nanomedicine.Prostate Cancer Foundation (Award in Nanotherapeutics)MIT-Harvard Center for Cancer Nanotechnology Excellence (U54-CA151884)National Heart, Lung, and Blood Institute (Programs of Excellence in Nanotechnology (HHSN268201000045C))National Science Foundation (U.S.) (Graduate Research Fellowship

Examining the Lateral Displacement of HL60 Cells Rolling on Asymmetric P-Selectin Patterns

Author Manuscript 2011 July 4.The lateral displacement of cells orthogonal to a flow stream by rolling on asymmetrical receptor patterns presents a new opportunity for the label-free separation and analysis of cells. Understanding the nature of cell rolling trajectories on such substrates is necessary to the engineering of substrates and the design of devices for cell separation and analysis. Here, we investigate the statistical nature of cell rolling and the effect of pattern geometry and flow shear stress on cell rolling trajectories using micrometer-scale patterns of biomolecular receptors with well-defined edges. Leukemic myeloid HL60 cells expressing the PSGL-1 ligand were allowed to flow across a field of patterned lines fabricated using microcontact printing and functionalized with the P-selectin receptor, leveraging both the specific adhesion of this ligand−receptor pair and the asymmetry of the receptor pattern inclination angle with respect to the fluid shear flow direction (α = 5, 10, 15, and 20°). The effects of the fluid shear stress magnitude (τ = 0.5, 1, 1.5, and 2.0 dyn/cm[superscript 2]), α, and P-selectin incubation concentration were quantified in terms of the rolling velocity and edge tracking length. Rolling cells tracked along the inclined edges of the patterned lines before detaching and reattaching on another line. The detachment of rolling cells after tracking along the edge was consistent with a Poisson process of history-independent interactions. Increasing the edge inclination angle decreased the edge tracking length in an exponential manner, contrary to the shear stress magnitude and P-selectin incubation concentration, which did not have a significant effect. On the basis of these experimental data, we constructed an empirical model that predicted the occurrence of the maximum lateral displacement at an edge angle of 7.5°. We also used these findings to construct a Monte Carlo simulation for the prediction of rolling trajectories of HL60 cells on P-selectin-patterned substrates with a specified edge inclination angle. The prediction of lateral displacement in the range of 200 μm within a 1 cm separation length supports the feasibility of label-free cell separation via asymmetric receptor patterns in microfluidic devices.Deshpande Center for Technological InnovationNational Science Foundation (U.S.) (CAREER Award 0952493)National Institutes of Health (U.S.) (Grant DE019191)National Institutes of Health (U.S.) (Grant HL095722)National Institutes of Health (U.S.) (Grant HL097172)American Heart Association (Grant 0970178N

Engineered mesenchymal stem cells with self-assembled vesicles for systemic cell targeting

Cell therapy has the potential to impact the quality of life of suffering patients. Systemic infusion is a convenient method of cell delivery; however, the efficiency of engraftment presents a major challenge. It has been shown that modification of the cell surface with adhesion ligands is a viable approach to improve cell homing, yet current methods including genetic modification suffer potential safety concerns, are practically complex and are unable to accommodate a wide variety of homing ligands or are not amendable to multiple cell types. We report herein a facile and generic approach to transiently engineer the cell surface using lipid vesicles to present biomolecular ligands that promote cell rolling, one of the first steps in the homing process. Specifically, we demonstrated that lipid vesicles rapidly fuse with the cell membrane to introduce biotin moieties on the cell surface that can subsequently conjugate streptavidin and potentially any biotinylated homing ligand. Given that cell rolling is a pre-requisite to firm adhesion for systemic cell homing, we examined the potential of immobilizing sialyl Lewis X (SLeX) on mesenchymal stem cells (MSCs) to induce cell rolling on a P-selectin surface, under dynamic flow conditions. MSCs modified with SLeX exhibit significantly improved rolling interactions with a velocity of 8 μm/s as compared to 61 μm/s for unmodified MSCs at a shear stress of 0.5 dyn/cm[superscript 2]. The cell surface modification does not impact the phenotype of the MSCs including their viability and multi-lineage differentiation potential. These results show that the transitory modification of cell surfaces with lipid vesicles can be used to efficiently immobilize adhesion ligands and potentially target systemically administered cells to the site of inflammation.American Heart Association (Grant 0970178N)National Institutes of Health (U.S.) (Grant DE019191

More than magnetic isolation: Dynabeads as strong Raman reporters towards simultaneous capture and identification of targets

Dynabeads are superparamagnetic particles used for immunomagnetic purification of cells and biomolecules. Post-capture, however, target identification relies on tedious culturing, fluorescence staining and/or target amplification. Raman spectroscopy presents a rapid detection alternative, but current implementations target cells themselves with weak Raman signals. We present antibody-coated Dynabeads as strong Raman reporter labels whose effect can be considered a Raman parallel of immunofluorescent probes. Recent developments in techniques for separating target-bound Dynabeads from unbound Dynabeads makes such an implementation feasible. We deploy Dynabeads anti-Salmonella to bind and identify Salmonella enterica, a major foodborne pathogen. Dynabeads present signature peaks at 1000 and 1600 1/cm from aliphatic and aromatic C-C stretching of polystyrene, and 1350 1/cm and 1600 1/cm from amide, alpha-helix and beta-sheet of antibody coatings of the Fe2O3 core, confirmed with electron dispersive X-ray (EDX) imaging. Their Raman signature can be measured in dry and liquid samples even at single shot ~30 x 30-micrometer area imaging using 0.5 s, 7 mW laser acquisition with single and clustered beads providing a 44- and 68-fold larger Raman intensity compared to signature from cells. Higher polystyrene and antibody content in clusters yields to the larger signal intensity and conjugation to bacteria strengthens clustering as a bacterium can bind to more than one bead as observed via transmission electron microscopy (TEM). Our findings shed light on the intrinsic Raman reporter nature of Dynabeads, demonstrating their dual function for target isolation and detection without additional sample preparation, staining, or unique plasmonic substrate engineering, advancing their applications in heterogeneous samples like food, water, and blood.Comment: 35 pages, 19 figures, submitted to the Journal of Raman Spectroscop

Microfluidic Platform for Combinatorial Synthesis and Optimization of Targeted Nanoparticles for Cancer Therapy

Taking a nanoparticle (NP) from discovery to clinical translation has been slow compared to small molecules, in part by the lack of systems that enable their precise engineering and rapid optimization. In this work we have developed a microfluidic platform for the rapid, combinatorial synthesis and optimization of NPs. The system takes in a number of NP precursors from which a library of NPs with varying size, surface charge, target ligand density, and drug load is produced in a reproducible manner. We rapidly synthesized 45 different formulations of poly(lactic-co-glycolic acid)-b-poly(ethylene glycol) NPs of different size and surface composition and screened and ranked the NPs for their ability to evade macrophage uptake in vitro. Comparison of the results to pharmacokinetic studies in vivo in mice revealed a correlation between in vitro screen and in vivo behavior. Next, we selected NP synthesis parameters that resulted in longer blood half-life and used the microfluidic platform to synthesize targeted NPs with varying targeting ligand density (using a model targeting ligand against cancer cells). We screened NPs in vitro against prostate cancer cells as well as macrophages, identifying one formulation that exhibited high uptake by cancer cells yet similar macrophage uptake compared to nontargeted NPs. In vivo, the selected targeted NPs showed a 3.5-fold increase in tumor accumulation in mice compared to nontargeted NPs. The developed microfluidic platform in this work represents a tool that could potentially accelerate the discovery and clinical translation of NPs.Prostate Cancer Foundation (Award in Nanotherapeutics)National Cancer Institute (U.S.) (Center of Cancer Nanotechnology Excellence at MIT-Harvard U54-CA151884National Heart, Lung, and Blood Institute (Programs of Excellence in Nanotechnology HHSN268201000045C)National Science Foundation (U.S.). Graduate Research FellowshipAmerican Society for Engineering Education. National Defense Science and Engineering Graduate FellowshipNational Cancer Institute (U.S.) (Center of Cancer Nanotechnology Excellence. Graduate Research Fellowship

A cell rolling cytometer reveals the correlation between mesenchymal stem cell dynamic adhesion and differentiation state

This communication presents quantitative studies of the dynamic adhesion behavior of mesenchymal stem cells (MSCs) enabled by the combination of cell-surface receptor–ligand interactions and three-dimensional hydrodynamic control by microtopography.National Institutes of Health (U.S.) (Grant HL-095722)National Institutes of Health (U.S.) (Grant HL-097172)National Science Foundation (U.S.) (CAREER Award 0952493)Korea (South). Ministry of Science, ICT and Future Planning (National Research Foundation of Korea. Pioneer Research Center Program 2013M3C1A3064777)National Research Foundation of Korea (Framework of International Cooperation Program 2013K2A1A2053078