Scaling of deterministic lateral displacement devices to a single column of bumping obstacles.

We describe a deterministic lateral displacement (DLD) for particle separation with only a single column of bumping features. The bifurcation of fluid streams at obstacles is not set by the "tilt" of columns with respect to macroscopic current flow, but rather by the fluidic resistances for lateral flow at each obstacle. With one column of 14 bumping features and corresponding inlet/outlet channels, the single-column DLD can separate particles with diameters of 4.8 µm and 9.9 µm at 30 µL min-1, with an area of only 0.37 mm × 1.5 mm (0.55 mm2). The large-cell output contains over 99% of the 9.9 µm particles and only 0.2% of the 4.8 m particles. The throughput per area of 54 µL min-1 per mm2 represents a 10× increase over previous selective harvesting reports for microfluidic devices in a similar particle size range.

An in vitro tumor swamp model of heterogeneous cellular and chemotherapeutic landscapes.

The heterogenous, highly metabolic stressed, poorly irrigated, solid tumor microenvironment - the tumor swamp - is widely recognized to play an important role in cancer progression as well as the development of therapeutic resistance. It is thus important to create realistic in vitro models within the therapeutic pipeline that can recapitulate the fundamental stress features of the tumor swamp. Here we describe a microfluidic system which generates a chemical gradient within connected microenvironments achieved through a static diffusion mechanism rather than active pumping. We show that the gradient can be stably maintained for over a week. Due to the accessibility and simplicity of the experimental platform, the system allows for not only well-controlled continuous studies of the interactions among various cell types at single-cell resolution, but also parallel experimentation for time-resolved downstream cellular assays on the time scale of weeks. This approach enables simple, compact implementation and is compatible with existing 6-well imaging technology for simultaneous experiments. As a proof-of-concept, we report the co-culture of a human bone marrow stromal cell line and a bone-metastatic prostate cancer cell line using the presented device, revealing on the same chip a transition in cancer cell survival as a function of drug concentration on the population level while exhibiting an enrichment of poly-aneuploid cancer cells (PACCs) as an evolutionary consequence of high stress. The device allows for the quantitative study of cancer cell dynamics on a stress landscape by real-time monitoring of various cell types with considerable experimental throughput.

Large-Area Resistive Strain Sensing Sheet for Structural Health Monitoring.

Damage significantly influences response of a strain sensor only if it occurs in the proximity of the sensor. Thus, two-dimensional (2D) sensing sheets covering large areas offer reliable early-stage damage detection for structural health monitoring (SHM) applications. This paper presents a scalable sensing sheet design consisting of a dense array of thin-film resistive strain sensors. The sensing sheet is fabricated using flexible printed circuit board (Flex-PCB) manufacturing process which enables low-cost and high-volume sensors that can cover large areas. The lab tests on an aluminum beam showed the sheet has a gauge factor of 2.1 and has a low drift of 1.5 µ ϵ / d a y . The field test on a pedestrian bridge showed the sheet is sensitive enough to track strain induced by the bridge's temperature variations. The strain measured by the sheet had a root-mean-square (RMS) error of 7 µ ϵ r m s compared to a reference strain on the surface, extrapolated from fiber-optic sensors embedded within the bridge structure. The field tests on an existing crack showed that the sensing sheet can track the early-stage damage growth, where it sensed 600 µ ϵ peak strain, whereas the nearby sensors on a damage-free surface did not observe significant strain change.

Generation of Heterogeneous Drug Gradients Across Cancer Populations on a Microfluidic Evolution Accelerator for Real-Time Observation.

Conventional cell culture remains the most frequently used preclinical model, despite its proven limited ability to predict clinical results in cancer. Microfluidic cancer-on-chip models have been proposed to bridge the gap between the oversimplified conventional 2D cultures and more complicated animal models, which have limited ability to produce reliable and reproducible quantitative results. Here, we present a microfluidic cancer-on-chip model that reproduces key components of a complex tumor microenvironment in a comprehensive manner, yet is simple enough to provide robust quantitative descriptions of cancer dynamics. This microfluidic cancer-on-chip model, the "Evolution Accelerator," breaks down a large population of cancer cells into an interconnected array of tumor microenvironments while generating a heterogeneous chemotherapeutic stress landscape. The progression and the evolutionary dynamics of cancer in response to drug gradient can be monitored for weeks in real time, and numerous downstream experiments can be performed complementary to the time-lapse images taken through the course of the experiments.

Hybrid LAE-CMOS Force-Sensing System Employing TFT-Based Compressed Sensing for Scalability of Tactile Sensing Skins.

Tactile sensing requires form-fitting and dense sensor arrays over large-areas. Hybrid systems, combining Large-Area Electronics (LAE) and silicon-CMOS ICs to respectively provide diverse sensing and high-performance computation/control, enable a platform for such sensing. A key challenge is that hybrid systems require a large number of interfaces between the LAE and CMOS domains, particularly as the number of sensors scales. This paper presents an architecture that exploits the attribute of signal sparsity, commonly exhibited in large-scale tactile-sensing applications, to reduce the interfacing complexity to a level set by the sparsity rather than the number of sensors. This enhances scalability compared to sequential-scanning and active-matrix approaches. The architecture implements compressed sensing via thin-film-transistor (TFT) switches, and is demonstrated in a force-sensing system with 20 force sensors, a TFT die (with 161 ZnO TFTs) per sensor, and a custom CMOS IC for system readout and control. Acquisition error of 0.7 k[Formula: see text] is achieved over a 100 k Ω-20 k Ω sensing range, at energy and rate of 2.46  µ J/frame and 31 fps.

The role of heterogeneous environment and docetaxel gradient in the emergence of polyploid, mesenchymal and resistant prostate cancer cells.

The ability of a population of PC3 prostate epithelial cancer cells to become resistant to docetaxel therapy and progress to a mesenchymal state remains a fundamental problem. The progression towards resistance is difficult to directly study in heterogeneous ecological environments such as tumors. In this work, we use a micro-fabricated "evolution accelerator" environment to create a complex heterogeneous yet controllable in-vitro environment with a spatially-varying drug concentration. With such a structure we observe the rapid emergence of a surprisingly large number of polyploid giant cancer cells (PGCCs) in regions of very high drug concentration, which does not occur in conventional cell culture of uniform concentration. This emergence of PGCCs in a high drug environment is due to migration of diploid epithelial cells from regions of low drug concentration, where they proliferate, to regions of high drug concentration, where they rapidly convert to PGCCs. Such a mechanism can only occur in spatially-varying rather than homogeneous environments. Further, PGCCs exhibit increased expression of the mesenchymal marker ZEB1 in the same high-drug regions where they are formed, suggesting the possible induction of an epithelial to mesenchymal transition (EMT) in these cells. This is consistent with prior work suggesting the PGCC cells are mediators of resistance in response to chemotherapeutic stress. Taken together, this work shows the key role of spatial heterogeneity and the migration of proliferative diploid cells to form PGCCs as a survival strategy for the cancer population, with implications for new therapies.

Strain Transfer for Optimal Performance of Sensing Sheet.

Sensing sheets based on Large Area Electronics (LAE) and Integrated Circuits (ICs) are novel sensors designed to enable reliable early-stage detection of local unusual structural behaviors. Such a device consists of a dense array of strain sensors, patterned onto a flexible polyimide substrate along with associated electronics. Previous tests performed on steel specimens equipped with sensing sheet prototypes and subjected to fatigue cracking pointed to a potential issue: individual sensors that were on or near a crack would immediately be damaged by the crack, thereby rendering them useless in assessing the size of the crack opening or to monitor future crack growth. In these tests, a stiff adhesive was used to bond the sensing sheet prototype to the steel specimen. Such an adhesive provided excellent strain transfer, but it also caused premature failure of individual sensors within the sheet. Therefore, the aim of this paper is to identify an alternative adhesive that survives minor damage, yet provides strain transfer that is sufficient for reliable early-stage crack detection. A sensor sheet prototype is then calibrated for use with the selected adhesive.

Cancer dormancy and criticality from a game theory perspective.

BACKGROUND: The physics of cancer dormancy, the time between initial cancer treatment and re-emergence after a protracted period, is a puzzle. Cancer cells interact with host cells via complex, non-linear population dynamics, which can lead to very non-intuitive but perhaps deterministic and understandable progression dynamics of cancer and dormancy. RESULTS: We explore here the dynamics of host-cancer cell populations in the presence of (1) payoffs gradients and (2) perturbations due to cell migration. CONCLUSIONS: We determine to what extent the time-dependence of the populations can be quantitively understood in spite of the underlying complexity of the individual agents and model the phenomena of dormancy.

Deterministic Lateral Displacement: The Next-Generation CAR T-Cell Processing?

Reliable cell recovery and expansion are fundamental to the successful scale-up of chimeric antigen receptor (CAR) T cells or any therapeutic cell-manufacturing process. Here, we extend our previous work in whole blood by manufacturing a highly parallel deterministic lateral displacement (DLD) device incorporating diamond microposts and moving into processing, for the first time, apheresis blood products. This study demonstrates key metrics of cell recovery (80%) and platelet depletion (87%), and it shows that DLD T-cell preparations have high conversion to the T-central memory phenotype and expand well in culture, resulting in twofold greater central memory cells compared to Ficoll-Hypaque (Ficoll) and direct magnetic approaches. In addition, all samples processed by DLD converted to a majority T-central memory phenotype and did so with less variation, in stark contrast to Ficoll and direct magnetic prepared samples, which had partial conversion among all donors (<50%). This initial comparison of T-cell function infers that cells prepared via DLD may have a desirable bias, generating significant potential benefits for downstream cell processing. DLD processing provides a path to develop a simple closed system that can be automated while simultaneously addressing multiple steps when there is potential for human error, microbial contamination, and other current technical challenges associated with the manufacture of therapeutic cells.

On-chip cell labelling and washing by capture and release using microfluidic trap arrays.

Flow cytometry analysis requires a large amount of isolated, labelled, and purified cells for accurate results. To address the demand for a large quantity of cells prepared in a timely manner, we describe a novel microfluidic trap structure array for on-chip cell labelling, such as intracellular and extracellular labelling, and subsequent washing and release of cells. Each device contains [Formula: see text] trap structures, which made the preparation of large numbers of cells [Formula: see text] possible. The structure has a streamlined shape, which minimizes clogging of cells in capture and release steps. The trap structure arrays are built and tested using leukocytes, with different load flow speeds, incubation times, and release flow speeds. ∼85% of cells are captured independent of the input flow speed. The release efficiency depends on the incubation time, with over ∼80% of captured cells released for up to 20 min incubation, and on-chip labelling and washing with STYO13 are demonstrated. Qualitative models are developed as guidance for designing the proposed trap structure and to explain the increased performance over previous approaches.

Epithelial and mesenchymal prostate cancer cell population dynamics on a complex drug landscape.

We have improved our microfluidic cell culture device that generates an in vitro landscape of stress heterogeneity. We now can do continuous observations of different cancer cell lines and carry out downstream analysis of cell phenotype as a function of position on the stress landscape. We use this technology to probe adaption and evolution dynamics in prostate cancer cell metapopulations under a stress landscape of a chemotherapeutic drug (docetaxel). The utility of this approach is highlighted by analysis of heterogenous prostate cancer cell motility changes as a function of position in the stress landscape. Because the technology presented here is easily adapted to a standard epifluorescence microscope it has the potential for broad application in preclinical drug development and assays of likely drug efficacy.

Automated leukocyte processing by microfluidic deterministic lateral displacement.

We previously developed a Deterministic Lateral Displacement (DLD) microfluidic method in silicon to separate cells of various sizes from blood (Davis et al., Proc Natl Acad Sci 2006;103:14779-14784; Huang et al., Science 2004;304:987-990). Here, we present the reduction-to-practice of this technology with a commercially produced, high precision plastic microfluidic chip-based device designed for automated preparation of human leukocytes (white blood cells; WBCs) for flow cytometry, without centrifugation or manual handling of samples. After a human blood sample was incubated with fluorochrome-conjugated monoclonal antibodies (mAbs), the mixture was input to a DLD microfluidic chip (microchip) where it was driven through a micropost array designed to deflect WBCs via DLD on the basis of cell size from the Input flow stream into a buffer stream, thus separating WBCs and any larger cells from smaller cells and particles and washing them simultaneously. We developed a microfluidic cell processing protocol that recovered 88% (average) of input WBCs and removed 99.985% (average) of Input erythrocytes (red blood cells) and >99% of unbound mAb in 18 min (average). Flow cytometric evaluation of the microchip Product, with no further processing, lysis or centrifugation, revealed excellent forward and side light scattering and fluorescence characteristics of immunolabeled WBCs. These results indicate that cost-effective plastic DLD microchips can speed and automate leukocyte processing for high quality flow cytometry analysis, and suggest their utility for multiple other research and clinical applications involving enrichment or depletion of common or rare cell types from blood or tissue samples. © 2016 International Society for Advancement of Cytometry.

Low-Temperature Synthesis of a TiO2/Si Heterojunction.

The classical SiO2/Si interface, which is the basis of integrated circuit technology, is prepared by thermal oxidation followed by high temperature (>800 °C) annealing. Here we show that an interface synthesized between titanium dioxide (TiO2) and hydrogen-terminated silicon (H:Si) is a highly efficient solar cell heterojunction that can be prepared under typical laboratory conditions from a simple organometallic precursor. A thin film of TiO2 is grown on the surface of H:Si through a sequence of vapor deposition of titanium tetra(tert-butoxide) (1) and heating to 100 °C. The TiO2 film serves as a hole-blocking layer in a TiO2/Si heterojunction solar cell. Further heating to 250 °C and then treating with a dilute solution of 1 yields a hole surface recombination velocity of 16 cm/s, which is comparable to the best values reported for the classical SiO2/Si interface. The outstanding performance of this heterojunction is attributed to Si-O-Ti bonding at the TiO2/Si interface, which was probed by angle-resolved X-ray photoelectron spectroscopy. Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) showed that Si-H bonds remain even after annealing at 250 °C. The ease and scalability of the synthetic route employed and the quality of the interface it provides suggest that this surface chemistry has the potential to enable fundamentally new, efficient silicon solar cell devices.

Microfluidic chemical processing with on-chip washing by deterministic lateral displacement arrays with separator walls.

We describe a microfluidic device for on-chip chemical processing, such as staining, and subsequent washing of cells. The paper introduces "separator walls" to increase the on-chip incubation time and to improve the quality of washing. Cells of interest are concentrated into a treatment stream of chemical reagents at the first separator wall for extended on-chip incubation without causing excess contamination at the output due to diffusion of the unreacted treatment chemicals, and then are directed to the washing stream before final collections. The second separator wall further reduces the output contamination from diffusion to the washing stream. With this approach, we demonstrate on-chip leukocyte staining with Rhodamine 6G and washing. The results suggest that other conventional biological and analytical processes could be replaced by the proposed device.

Ancient hot and cold genes and chemotherapy resistance emergence.

We use a microfabricated ecology with a doxorubicin gradient and population fragmentation to produce a strong Darwinian selective pressure that drives forward the rapid emergence of doxorubicin resistance in multiple myeloma (MM) cancer cells. RNA sequencing of the resistant cells was used to examine (i) emergence of genes with high de novo substitution densities (i.e., hot genes) and (ii) genes never substituted (i.e., cold genes). The set of cold genes, which were 21% of the genes sequenced, were further winnowed down by examining excess expression levels. Both the most highly substituted genes and the most highly expressed never-substituted genes were biased in age toward the most ancient of genes. This would support the model that cancer represents a revision back to ancient forms of life adapted to high fitness under extreme stress, and suggests that these ancient genes may be targets for cancer therapy.

Concentrating genomic length DNA in a microfabricated array.

We demonstrate that a microfabricated bump array can concentrate genomic-length DNA molecules efficiently at continuous, high flow velocities, up to 40 µm/s, if the single-molecule DNA globule has a sufficiently large shear modulus. Increase in the shear modulus is accomplished by compacting the DNA molecules to minimal coil size using polyethylene glycol (PEG) derived depletion forces. We map out the sweet spot, where concentration occurs, as a function of PEG concentration and flow speed using a combination of theoretical analysis and experiment. Purification of DNA from enzymatic reactions for next-generation DNA-sequencing libraries will be an important application of this development.

Inhibition of clot formation in deterministic lateral displacement arrays for processing large volumes of blood for rare cell capture.

Microfluidic deterministic lateral displacement (DLD) arrays have been applied for fractionation and analysis of cells in quantities of ~100 µL of blood, with processing of larger quantities limited by clogging in the chip. In this paper, we (i) demonstrate that this clogging phenomenon is due to conventional platelet-driven clot formation, (ii) identify and inhibit the two dominant biological mechanisms driving this process, and (iii) characterize how further reductions in clot formation can be achieved through higher flow rates and blood dilution. Following from these three advances, we demonstrate processing of 14 mL equivalent volume of undiluted whole blood through a single DLD array in 38 minutes to harvest PC3 cancer cells with ~86% yield. It is possible to fit more than 10 such DLD arrays on a single chip, which would then provide the capability to process well over 100 mL of undiluted whole blood on a single chip in less than one hour.

Improved efficiency of hybrid organic photovoltaics by pulsed laser sintering of silver nanowire network transparent electrode.

In this Research Article, we demonstrate pulsed laser processing of a silver nanowire network transparent conductor on top of an otherwise complete solar cell. The macroscopic pulsed laser irradiation serves to sinter nanowire-nanowire junctions on the nanoscale, leading to a much more conductive electrode. We fabricate hybrid silicon/organic heterojunction photovoltaic devices, which have ITO-free, solution processed, and laser processed transparent electrodes. Furthermore, devices which have high resistive losses show up to a 35% increase in power conversion efficiency after laser processing. We perform this study over a range of laser fluences, and a range of nanowire area coverage to investigate the sintering mechanism of nanowires inside of a device stack. The increase in device performance is modeled using a simple photovoltaic diode approach and compares favorably to the experimental data.

Ratchets in hydrodynamic flow: more than waterwheels.

The transport of objects in microfluidic arrays of obstacles is a surprisingly rich area of physics and statistical mechanics. Tom Duke's mastery of these areas had a major impact in the development of biotechnology which uses these ideas at an increasing scale. We first review how biological objects are transported in fluids at low Reynolds numbers, including a discussion of electrophoresis, then concentrate on the separation of objects in asymmetric arrays, sometimes called Brownian ratchets when diffusional symmetry is broken by the structures. We move beyond this to what are called deterministic arrays where non-hydrodynamic forces in asymmetric arrays allow for extraordinary separation, and we look to the future of using these unusual arrays at the nanoscale and at the hundreds of micrometre scale. The emphasis is on how the original ideas of Tom Duke drove this work forward.

You cannot tell a book by looking at the cover: Cryptic complexity in bacterial evolution.

Do genetically closely related organisms under identical, but strong selection pressure converge to a common resistant genotype or will they diverge to different genomic solutions? This question gets at the heart of how rough is the fitness landscape in the local vicinity of two closely related strains under stress. We chose a Growth Advantage in Stationary Phase (GASP) E scherichia coli strain to address this question because the GASP strain has very similar fitness to the wild-type (WT) strain in the absence of metabolic stress but in the presence of metabolic stress continues to divide and does not enter into stationary phase. We find that under strong antibiotic selection pressure by the fluoroquinolone antibiotic ciprofloxacin in a complex ecology that the GASP strain rapidly evolves in under 20 h missense mutation in gyrA only 2 amino acids removed from the WT strain indicating a convergent solution, yet does not evolve the other 3 mutations of the WT strain. Further the GASP strain evolves a prophage e14 excision which completely inhibits biofilm formation in the mutant strain, revealing the hidden complexity of E. coli evolution to antibiotics as a function of selection pressure. We conclude that there is a cryptic roughness to fitness landscapes in the absence of stress.