The Personalized Health Care Environment: How in vitro Diagnostics, Mobile Health and Medical Devices will Converge to Improve Health Care
Alan Wright, Chief Medical Officer, Roche Diagnostics Corporation, United States of America

Digital Microfluidics: A Platform Whose Time Has Come
Aaron Wheeler, Canada Research Chair of Bioanalytical Chemistry, University of Toronto, Canada

Digital microfluidics is a fluid-handling technique in which droplets are manipulated by electrostatic forces on an array of electrodes coated with a hydrophobic insulator. In this talk, I will present recent results from my group’s work with digital microfluidics, with an emphasis on the topics covered in this unique venue. Specifically, I will demonstrate how digital microfluidics is particularly well-suited for Lab on a Chip applications, given its ability to automate diverse laboratory processes on a generic, programmable platform. Likewise, I will report on our work using digital microfluidics for Point of Care Diagnostics and Global Health, reporting on the results of a field trial for measles and rubella diagnostics at refugee sites in Kenya. Finally, I will describe how digital microfluidics is emerging as a useful tool for Single-Cell Analysis and for integration with Mass Spectrometry to answer questions about cell heterogeneity and cell-cell communication. Through these examples, I will make the case that digital microfluidics is emerging as a useful new tool for the next generation of analytical techniques, across a wide range of applications.

Microfluidics and Sensors: New Tools for Real-Time Clinical Monitoring
Martyn Boutelle, Professor of Biomedical Sensors Engineering, Imperial College London, United Kingdom

A goal for modern medicine is to protect vulnerable tissue by monitoring the patterns of changing physical,  electrical and chemical changes taking place in tissue - ‘multimodal monitoring’. Clinicians hope such information will allows treatments to be guided and ultimately controlled based on the measured signals. Microfluidic lab-on-chip devices coupled to tissue sampling using microdialysis provide an important new way for measuring real-time chemical changes as the low volume flow rates  of microdialysis probes are ideally matched to the length scales of microfluidic devices. Concentrations of key biomarker molecules can then be determined continuously using either optically or electrochemically (using amperometric, and potentiometic sensors).  Wireless devices allow analysis to take place close to the patient. Droplet-based microfluidics, by digitizing the dialysis stream into discrete low volume samples, both minimizes dispersion allowing very rapid concentration changes to be measured, and allows rapid transport of samples between patient and analysis chip. This talk will overview successful design, optimization, automatic-calibration and use of both continuous flow and droplet-based microfluidic analysis systems for real-time clinical monitoring, using clinical examples from our recent work.

Microfluidics for Personalized Oncology
Chwee Teck Lim, NUS Society Chair Professor, Department of Biomedical Engineering, Institute for Health Innovation & Technology (iHealthtech), Mechanobiology Institute, National University of Singapore, Singapore

Cancer is a disease that can be characterised by the heterogeneity of the tumor cells.  Such heterogeneity has led to a major hindrance in cancer classification, diagnosis and treatment. Recently, single cell analysis has become an important approach to detect variations in genetic, proteomic and molecular expressions in individual cancer cells.  Here, we developed a suite of microfluidic biochips that makes use of cell mechanics principle to not only isolate, but also enable single cell analysis of circulating tumor cells (CTCs) derived from the peripheral blood of cancer patients.  These include detecting the proteolytic capability of each CTC to obtain insights into their invasiveness as well as identify unique mutations with the aim of improving treatment.  It is hope that this approach will lead to better personalized treatment of cancer patients.

High-Performance Rapid Diagnostic Tests
Bernhard Weigl, Director, Center for In-Vitro Diagnostics, Intellectual Ventures/Global Good-Bill Gates Venture Fund, United States of America

Lateral flow and similar rapid diagnostic assays (LFAs) are easy to use and manufacture, low cost, rapid, require little or no equipment to operate, and do not need to be refrigerated. However, they are generally not considered to be very sensitive or able to provide a quantitative result. Our group believes that this lack of sensitivity is not a fundamental property of LFAs but rather a consequence of the way they are developed, manufactured, and marketed. Historically, most lateral flow tests were developed and optimized by relatively small manufacturers with limited R&D capabilities and budgets, and were generally used only for analytical targets prevalent at high concentration in patient’s samples that were relatively easy to measure. In contrast, our group’s mission is to develop LFA-based assays for use in global health applications that are as sensitive as the best conventional diagnostic assays (in some cases even better) while retaining all their cost, simplicity, and usability advantages.

Technologies for Personalizing Cancer Immunotherapies
James Heath, Elizabeth W. Gilloon Professor of Chemistry, California Institute of Technology (CalTech), United States of America

Cancer immunotherapy, which has taken virtually all aspects of oncology by storm over the past few years, is based upon using  cellular or molecular therapies to promote tumor cell/immune cell interactions.  At the heart of this therapy are the T cells that actually do the tumor cell killing, and the tumor antigens that are recognized by those T cells.  Recent work has shown that neoantigens play critical roles in many immunotherapy successes.  Neoantigens are tumor antigens that are fragments of mutated proteins expressed by the cancer cells, and contain those point mutations.  They are presented in the clefts of major histocompatibility molecules (MHCs) by many of the cells in the tumor, where they may be recognized by neoantigen-specific T cell populations.  In this presentation, I will discuss how various micro and nanotechnologies are being harnessed to identify, for a given patient which neoantigens are actively recruiting T cells into the tumor, and to carry out a deep molecular analysis of those neoantigen-specific T cells.  I will further discuss how that information can then be harnessed for personalized cancer immunotherapies in the form of neoantigen-based vaccines, or engineered T cell receptor adoptive cell transfer therapies.

Centrifugal Microfluidics for Biomedical Applications
Yoon-Kyoung Cho, Professor, Biomedical Engineering, Ulsan National Institute of Science & Technology; Group leader, IBS; FRSC, Fellow of Royal Society of Chemistry, Korea South

In this presentation, we will discuss our on-going research on “Lab-on-a-disc”, which applies centrifugal force to pump fluid for biochemical analysis. It is advantageous because of the capability to integrate and automate the analysis protocols into a disc-shaped device with simple, size-reduced, and cost-efficient instrumentation. We report various examples of fully integrated "lab-on-a-disc" for biomedical applications such as pathogen specific DNA extraction to test infectious diseases, multiplex enzyme-linked immunosorbent assay (ELISA), and isolation and analysis of circulating tumor cells starting from whole blood. Integration with centrifugal microfluidic technology allows us precise control of fluids while also reducing the expensive reagent consumption, the required analysis time and possible handling errors. We believe the presented result will not only improve the performance of the point-of-care-diagnostic devices but also potentially have great impact on global healthcare.

A Billion-Droplet AC Electrospray Digital PCR Platform for Large-Dynamic Range Nucleic Acid Quantification
Hsueh-Chia Chang, Bayer Professor of Chemical and Biomolecular Engineering, University of Notre Dame, Interim Chief Technology Officer, Aopia Biosciences, United States of America

An AC-electrospray technology is shown to be able to generate 1 billion femto-liter (micron diameter) aqueous drops in immiscible silicone oil in less than 20 minutes.  Unlike DC sprays, which tend to suffer from dielectric breakdown in liquid, and droplet generation technologies based on hydrodynamic shear, which can only generate pico-liter drops, we have shown that a properly tuned AC field can entrain low-mobility anions at a meniscus and the Coulombic repulsion among such entrained ions can deform the meniscus into unique 11 degree AC cones that are quite distinct from and much stable than DC cones (Phys Rev Lett, 101, 204501(2008); 109, 224301(2012); LabChip, 15, 1656(2015)).   The sharper AC cones can generate 1 million femto-liter drops per second.  The larger drop number and concentration (nM) allow us to get significantly better accuracy (without using Poisson statistics), dynamic range (8 decades) and reduction of inhibition and interfering effects in heterogeneous media.  We have fabricated an integrated AC electrospray chip with PCR and imaging modules and have favorably compared the performance of this new droplet digital PCR platform against qPCR. The new AC droplet digital PCR platform is, however, a low-cost turn-key technology that requires no or minimum sample pretreatment.

Cost-Effective Microfluidic Platform for Point-of-Care Diagnostics and Various Life-Science Applications
Alexander Govyadinov, Senior Technologist, HP Incorporated, United States of America

Recently, there is a lot of interest in microfluidic lab-on-a chip application for life science, forensic, point-of-care, molecular-diagnostic, other in-vitro-diagnostic, organs-on-a-chip, environmental and multiple others applications. Different scientific and commercial organizations explore a multitude of material sets and operational principles to forge microfluidic devices. Simultaneously, inkjet industry utilizes well established materials and principles of operation for complicated microfluidic systems developed for precision dispensing and manipulation of droplets with pico-liter accuracy on a massively parallel scale. The presentation describes our recent progress of low cost microfluidic platform development utilizing materials and processes developed for low-cost thermal inkjet business. The concept repurposes well established inkjet processes, microfluidic components and jetting elements for pumping, mixing, valving, fluid transport, sensing and other critical functions of complex integrated microfluidic systems.  This presentation describes operating principles of microfluidic elements, examples of their integration in functional devices and discusses inkjet technology potential for broad range of microfluidic applications.

The Art of 3D-Printing Biocompatible Microfluidics
Albert Folch, Professor of Bioengineering, University of Washington, United States of America

The vast majority of microfluidic systems are presently built by replica-molding and bonding in elastomers (such as poly(dimethyl siloxane) (PDMS)) or in thermoplastics (such as poly(methyl methacrylate) (PMMA) or poly-styrene (PS)). However, biologists and clinicians typically do not have access to microfluidic technology because they do not have the engineering expertise or equipment required to fabricate and/or operate microfluidic devices. Furthermore, the present commercialization path for microfluidic devices is usually restricted to high-volume applications in order to recover the large investment needed to develop the plastic molding processes. We are developing microfluidic devices through stereolithography, a form of 3D printing, in order to make microfluidic technology readily available via the web to biomedical scientists. Most available SL resins do not have all the favorable physicochemical properties of the above-named plastics (e.g., biocompatibility, transparency, elasticity, and gas permeability), so the performance of SL-printed devices is still inferior to that of equivalent PDMS devices. Inspired by the success of hydrogel PEG-DA biocompatibility, we have developed microfluidic devices by SL in resins that share all the advantageous attributes of PDMS and thermoplastics so that we can 3D-print designs with comparable performance and biocompatibility to those that are presently molded.

Microfluidic Printing: From Combinatorial Drug Screening to Artificial Cell Assaying
Tingrui Pan, Professor, Department of Biomedical Engineering, University of California-Davis, United States of America

Microfluidic impact printing has been recently introduced, benefiting from the nature of simple device architecture, low cost, non-contamination, scalable multiplexability and high throughput.  In this talk, we will review this novel microfluidic-based droplet generation platform, utilizing modular microfluidic cartridges and expandable combinatorial printing capacity controlled by plug-and-play multiplexed actuators.  Such a customizable microfluidic printing system allows for ultrafine control of the droplet volume from picoliters (~10pL) to nanoliters (~100nL), a 10,000 fold variation. The high flexibility of droplet manipulations can be simply achieved by controlling the magnitude of actuation (e.g., driving voltage) and the waveform shape of actuation pulses, in addition to nozzle size restrictions. Detailed printing characterizations on these parameters have been conducted consecutively. A multichannel impact printing system has been prototyped and demonstrated to provide the functions of single-droplet jetting and droplet multiplexing as well as concentration gradient generation. Moreover, several enabling chemical and biological assays have been implemented and validated on this highly automated and flexible printing platform.  In brief, the microfluidic impact printing system could be of potential value to establishing multiplexed droplet assays for high-throughput life science researchers.

Nanopore Sequencing for Real-Time Pathogen Identification
Kamlesh Patel, R&D Advanced System Engineering and Deployment Manager, Sandia National Laboratories, United States of America

As recent outbreaks have shown, effective global health response to emergent infectious disease requires a rapidly deployable, universal diagnostic capability. We will present our ongoing work to develop a fieldable device for universal bacterial pathogen characterization based on nanopore DNA sequencing. Our approach leverages synthetic biofunctionalized nanopore structures to sense each nucleotide. We aim to create a man-portable platform by combining nanopore sequencing with advance microfluidic-based sample preparation methods for an amplification-free, universal sample prep to accomplish multiplexed, broad-spectrum pathogen and gene identification.

Polymer-based Nanosensors using Flight-Time Identification of Mononucleotides for Single-Molecule Sequencing
Steve Soper, Foundation Distinguished Professor, Director, Center of BioModular Multi-Scale System for Precision Medicine, The University of Kansas, United States of America

We are generating a single-molecule DNA sequencing platform that can acquire sequencing information with high accuracy. The technology employs high density arrays of nanosensors that read the identity of individual mononucleotides from their characteristic flight-time through a 2-dimensional (2D) nanochannel (~20 nm in width and depth; >100 µm in length) fabricated in a thermoplastic via nano-imprinting (NIL). The mononucleotides are generated from an intact DNA fragment using a highly processive exonuclease, which is covalently anchored to a plastic solid support contained within a bioreactor that sequentially feeds mononucleotides into the 2D nanochannel. The identity of the mononucleotides is deduced from a molecular-dependent flight-time through the 2D nanochannel. The flight time is read in a label-less fashion by measuring current transients induced a single mononucleotide when it travels through a constriction with molecular dimensions (<10 nm in diameter) that are poised at the input/output ends of the flight tube. In this presentation, our efforts on building these polymer nanosensors using NIL in thermoplastics will be discussed and the detection of single molecules using electrical transduction with their identity deduced from the associated flight time provided. Finally, information on the manipulation of single DNA molecules using nanofluidic circuits will be discussed that takes advantage of forming unique nano-scale features to shape electric fields for DNA manipulation and serves as the functional basis of the nanosensing platform.

Rapid and Ultra-sensitive Diagnostics Using Digital Detection
Weian Zhao, Associate Professor, Department of Biomedical Engineering, University of California-Irvine, United States of America

We will present our most recent droplet based digital detection platforms for rapid and sensitive detections, which could find potential applications at the point-of-care (POC).

Fractionation and Analysis of Nuclear versus Cytoplasmic Nucleic Acids from Single Cells
Juan Santiago, Charles Lee Powell Foundation Professor, Stanford University, United States of America

Single cell analyses (SCA) have become powerful tools in the study heterogeneous cell populations such as tumors and developing embryos.  However, fractionating and analyzing nuclear versus cytoplasmic fractions of nucleic acids remains a challenge as these fractions easily cross-contaminate.  We present a novel microfluidic system that can fractionate and deliver nucleic acid (NA) fractions from the nucleus (nNA) versus the cytoplasm (cNA) from single cells to independent downstream analyses.  Our technique leverages a selective electrical lysis which disrupts the cell’s (outer) cytoplasmic membrane, while leaving the nucleus relatively intact. We selectively extract, purify, and preconcentrate cNA using isotachophoresis (ITP).  The ITP-focused cNA and nNA-containing nucleus are separated by ITP and fractionated at a bifurcation downstream and then extracted for off chip analyses. We will present example applications of this fractionation including qPCR and next generation sequencing (NGS) analyses of cNA vs. nNA.  This will include preliminary NGS analyses of nuclear vs. cytoplasmic RNA fractions to analyze gene expression and splicing.  We hypothesize that the robust and precise nature of our electric field control is amenable to further automation to increase throughput while removing manuals steps.

Novel Strategies in Single Molecule Sensing
Joshua Edel, Professor, Imperial College London, United Kingdom

Analytical Sensors plays a crucial role in today’s highly demanding exploration and development of new detection strategies. Whether it be medicine, biochemistry, bioengineering, or analytical chemistry the goals are essentially the same: [1.] improve sensitivity, [2.] maximize throughput, [3.] and reduce the instrumental footprint. In order to address these key challenges, the analytical community has borrowed technologies and design philosophies which has been used by the semiconductor industry over the past 20 years. By doing so, key technological advances have been made which include the miniaturization of sensors and signal processing components which allows for the efficient detection of nanoscale object. One can imagine that by decreasing the dimensions of a sensor to a scale similar to that of a nanoscale object, the ultimate in sensitivity can potentially be achieved - the detection of single molecules. This talk highlights novel strategies for the detection of single molecules.

Biological Imaging Using Microfluidics and Electrochemistry
Charles Henry, Professor and Chair, Colorado State University, United States of America

Chemical gradients drive many processes in biology, ranging from nerve signal transduction to ovulation to cancer metastasis. At present, microscopy is the primary tool used to understand these gradients by imaging both the gradients and the resulting cell motion. Microscopy has provided many important breakthroughs in our understanding of fundamental biology, but is limited due to the need to incorporate fluorescent molecules into biological systems through either labeling or genetic manipulations. To better understand some processes, there is a need to develop tools that can measure chemical gradient formation in biological systems that do not require fluorescent modification of the targets, can be multiplexed to measure more than one molecule, and are compatible with a variety of biological sample types, including in vitro cell cultures and ex vivo tissue slices. In response to this need, we have developed a high-density electrode array containing 8,192 individual electrodes to image release of electrochemically active metabolites like nitric oxide and norepinephrine from live tissue slices. The electrode array has a resolution of 30 µm and covers and area of 2 mm by 2 mm, easily large enough to image release of neurotransmitters across an ex vivo tissue slice from a mouse model. In this talk, electrochemical characterization of this system will be discussed first using simple chemical models. Then use of the system in combination with microfluidics, for imaging spatiotemporal resolution of neurotransmitter release at the organ scale will be presented. Use of microfluidics to deliver stimulants that induce changes in release profile will be shown.

Human “Body-on-a-Chip” Devices as Tool to Improve Drug Development
Michael Shuler, Samuel B. Eckert Professor of Engineering, Cornell University, President Hesperos, Inc., United States of America

Alternatives or supplements to the use of animals in preclinical drug development that better mimic human response should reduce costs and increase the number of FDA approved drugs at the end of clinical trials. We have constructed micro-physiological (or “Body-on-a-Chip”) devices constructed from a combination of human tissue engineered constructs, micro-fabricated devices and physiologically based pharmacokinetic (PBPK) models. These human surrogates are constructed on a low cost, robust “pumpless” platform. In addition to measuring viability and metabolic responses, we can measure functional outputs such as electrical activity and force generation using integrated sensors (in collaboration with J. Hickman, University of Central Florida). We will focus our discussion on development of key organ modules and their integration with each other to form a model of the human body.

Microraft Array Platform for the Selection of Lymphocytes Based on Target-Cell Killing
Nancy Allbritton, Frank and Julie Jungers Dean of the College of Engineering and Professor of Bioengineering, University of Washington in Seattle, United States of America

Adoptive cellular therapy (ACT) is an emerging therapeutic in which cytotoxic T lymphocytes (CTLs) that recognize tumor cell epitopes are introduced into patients providing immunity against the cancer cells. For ACT to succeed, CTLs with high tumor-killing efficiency must be identified, isolated, and characterized. Current technologies do not enable simultaneous assay of cell behavior or killing followed by recovery of the most efficient killer cells. A microraft array technology that measures the ability of individual T cells to lyse a population of target cells followed by sorting of living cells into a multi-well plate for expansion and characterization was developed.  The microraft array platform was combined with image processing and analysis algorithms to track and monitor killing assays over many hours. Automated cell collection was incorporated into the platform for facile cell collection from the array. As a proof of principle, human T cells directed against an influenza antigen were co-cultured with antigen presenting target cells on the microraft arrays. Target cell killing was measured by tracking the appearance of dead cells on each microraft over time. Microrafts with a single CTL demonstrating the greatest rate of target cell death were identified, cloned, and influenza-antigen reactivity confirmed. The platform is readily modified to measure the antigen-specific activity of individual cells within a bulk CTL culture or the cell heterogeneity within a population of gene-engineered T cells.