Lab-on-a-Chip and Microfluidics Europe 2021 Agenda

Microfluidics and Nanobiosensors for Enrichment and Detection of Biological Entities from Complex Matrices
Lorena Diéguez, Leader of the Medical Devices Research Group, INL- International Iberian Nanotechnology Laboratory, Portugal

Microfluidics presents numerous advantages for the handling of biological samples, as it provides careful control of fluids in the microscale. When it comes to the enrichment of biological entities, microfluidics has demonstrated superior sensitivity and enhanced recovery compared to traditional methods. Incorporating sensors, lab-on-a-chip technologies offer efficient characterization of biological entities from complex matrices, making microfluidics ideal for field applications, enabling high throughput, portability, and automation. In this talk, we present our work for microfluidic isolation and multiplex analysis of biological entities relevant for health, food and environment applications.

Microfluidics in Resource-Limited Settings
Nicole Pamme, Professor in Analytical Chemistry, Stockholm University, Sweden

Our research centers on the study of microfluidic lab-on-a-chip devices applied to environmental analysis, biomedical research and the synthesis of smart materials. Microfluidic devices offer the possibility for in-the-field and point-of-care analysis provided the devices are portable, require only minimal external instrumentation and little power and are robust. In our group, we are investigating a simple magnetic particle-based extraction method, IFAST (immiscible filtration based on surface tension), for pathogen isolation from clinical and environmental matrices in collaboration with researchers in South Africa and Kenya. Furthermore, as part of an EU-funded project (Sullied Sediments) we are developing paper microfluidic devices for on-site measurement of water quality markers such as phosphate. These are being tested by members of the general public who upload data via a custom-built up for data collection with high spatial and temporal resolution.

Optofluidic Imaging Flow Cytometry
Andrew J deMello, Professor of Biochemical Engineering & Institute Chair, ETH Zürich, Switzerland

I will present recent activities aimed at developing novel microfluidic imaging flow cytometers for blur-free cellular analysis at throughputs exceeding 100,000 cells per second. By combining passive (inertial or viscoelastic) focusing of cells in parallel microchannels with stroboscopic illumination, such chip-based cytometers are able to extract multi-colour fluorescence and bright-field images of single cells moving at high linear velocities. This in turn allows accurate sizing of individual cells, intracellular localization and analysis of heterogeneous cell suspensions. The methods are showcased through the rapid enumeration of apoptotic cells, high-throughput discrimination cell cycle phases and localization of p-bodies. Additionally, the optofluidic platforms can be integrated with weakly-supervised deep learning methods to perform rapid diagnosis of diseases, such as cutaneous T-cell lymphoma.

Droplet-based Microfluidics for Cancer Research: Application for Cancer Patient Follow-up
Valérie Taly, CNRS Research Director, Professor and Group leader Translational Research and Microfluidics, Université Paris Cité, France

Droplet-based microfluidics has led to the development of highly powerful tools with great potential in High-Throughput Screening where individual assays are compartmentalized within aqueous droplets acting as independent microreactors. Thanks to the combination of a decrease of assay volume and an increase of throughput, this technology goes beyond the capacities of conventional screening systems. Added to the flexibility and versatility of platform designs, such progresses in the manipulation of sub-nanoliter droplets has allowed to dramatically increase experimental level of control and precision. The presentation will aim at demonstrating through selected example, the great potential of this technology for biotechnology and cancer research. We will also show how by combining microfluidic systems and clinical advances in molecular diagnostic we have developed an original method to perform millions of single molecule PCR in parallel to detect and quantify a minority of target sequences in complex mixture of DNA with a sensitivity unreachable by conventional procedures. To demonstrate the pertinence of our procedures to overcome clinical oncology challenges, the results of clinical studies will be presented. In particular, applications of ddPCR to the follow-up of both advanced and localized digestive cancers will be presented. Finally, we will also show application of these technology for Covid19 patients.

Intestine on Chip for Normal Physiology and Disease Models
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

Organ-on-chips are miniaturized devices that arrange living cells to simulate functional subunits of tissues and organs. These microdevices provide exquisite control of tissue microenvironment for the investigation of organ-level physiology and disease. A 3D polarized epithelium using primary human or mouse gastrointestinal stem cells was developed to fully recapitulate gastrointestinal epithelial architecture and physiology. A planar monolayer comprised of stem/proliferative and differentiated primary cells is cultured on a shaped hydrogel scaffold with an array of crypt-like structures replicating the intestinal architecture. Imposition of chemical gradients across the crypt long axis yields a polarized epithelium with a stem-cell niche and differentiated cell zone. The stem cells proliferate, migrate and differentiate along the crypt axis as they do in vivo. A dense mucus layer is formed on the luminal epithelial surface that is impermeable to bacteria and acts a barrier to toxins. An oxygen gradient across the tissue mimic permits luminal culture of anaerobic bacteria while maintaining an oxygenated stem cell niche. This in vitro human colon crypt array replicates the architecture, luminal accessibility, tissue polarity, cell migration, and cellular responses of in vivo intestinal crypts. A thick impenetrable layer of mucus with biophysical parameters similar to that of a living human can be formed for epithelial cell-microbe studies. Intestinal biopsy samples can be used to populate these constructs to produce patient-specific tissues for personalized medicine and disease modeling. This bioanalytical platform is envisioned as a next-generation system for assay of microbiome-behavior, drug-delivery and toxin-interactions with the intestinal epithelia.

Isolation and Analysis of Liquid Biopsy Markers using Microfluidics and a Liquid Handling Robot
Steve Soper, Foundation Distinguished Professor, Director, Center of BioModular Multi-Scale System for Precision Medicine, The University of Kansas, United States of America

Liquid biopsies are generating great interest within the medical community due to the simplicity for securing important biomarkers to manage complex diseases, such as many of the cancer-related diseases. We are developing a suite of novel microfluidic devices that can process clinical samples directly and efficiently search for a variety of disease-associated liquid biopsy markers, such as circulating tumor cells (CTCs), cell free DNA (cfDNA), and extracellular vesicles (EVs). The microfluidic chips are made in plastics via injection molding and designed to optimize recovery and purity of each of the aforementioned biomarkers. The challenge with microfluidics is the need for attaching connector tubing to the microfluidic and using syringe pumps to fluidically operate the chips. We have developed liquid handling robots to automatically operate microchips and do so in a high throughput and automated fashion, appropriate for use in centralized clinical testing laboratories. The robots operate using pipet tips and do not require connector tubing reducing chip production costs. The robotic platforms were used along with specifically designed chips for analysis of each of the liquid biopsy markers. Examples of using the system for supporting large clinical studies will be discussed, such as CTCs for discovering new therapies for pancreatic cancer, cell free DNA genotyping for breast cancer, and EVs for patient stratification in ovarian and breast cancers.

Home Diagnostic Testing Before, During, and After the Era of COVID-19
Paul Yager, Professor, Department of Bioengineering, University of Washington, United States of America

For decades, testing of human samples for acute and chronic diseases has been in centralized laboratories where tests were carried out by trained technicians or by large robotic instruments capable of batch processing hundreds of tests.  While some home sampling had begun to be popular in the last decade (where samples were collected by individuals in the home, but those were mailed to a laboratory to carry out the testing), the only actual home testing was for pregnancy, HIV antibodies and blood glucose.  The COVID-19 pandemic, and the restriction of large portions of the world’s population to their homes has opened up new markets for many types of rapid home testing.  Since 1992 the Yager lab has focused on development of microfluidics for the analysis of biological fluids for use in low-cost point-of-care biomedical diagnostics for the developed and developing worlds.  Since 2008, the focus shifted to two-dimensional porous networks (“paper microfluidics") for ultra-low-cost instrument-free point-of-care pathogen identification.  Readout is often coupled optical imaging for quantitative analysis and data transmission; this has been under support of NIH, NSF, DARPA, DTRA.  In the last year the focus has been on developing rapid mutiplexed nucleic acid testing for respiratory pathogens for the home under support of WRF and the Emergent Ventures fund.

Sorting of Single Cells Based on Secreted Products Using Lab on a Particle Technology
Dino Di Carlo, Armond and Elena Hairapetian Chair in Engineering and Medicine, Professor and Vice Chair of Bioengineering, University of California-Los Angeles, United States of America

Sorting cells based on secreted products enables the discovery of antibodies, the development of cell lines producing recombinant products, and the selection of functional cells for cell therapies. However, approaches to rapidly analyze and sort viable cells based on secretions are not easily accessible. I will discuss nanovial technology that enables the isolation of individual cells in cavities of 3D-shaped hydrogel particles massively in parallel. These hydrogel particles, or nanovials, act as small wells to bind and accumulate secreted products at high concentrations, and enable washing and labeling steps. The nanovials and attached cells can then be sorted based on their secretions and their functions using standard FACS systems. This approach promises to democratize the ability to discover and manufacture drugs and cell therapies.

Designing a Bioreagent-Compatible Material for a 3D-Printed Molecular Design System
Noah Malmstadt, Professor, Mork Family Dept. of Chemical Engineering & Materials Science, University of Southern California, United States of America

While stereolithographic 3D printing (SLA) is a promising method for the rapid prototyping and manufacturing of microfluidic systems, the bioadhesive properties of cured SLA resins are poorly characterized. Adhesion of biomolecules to microfluidic channels is an issue in nearly all biological applications, but it becomes a particular problem in applications that require precise and reproducible control of reaction conditions. Over the past several years, we have deployed SLA-printed modular microfluidic components to automate the biochemical workflow of mRNA display. mRNA display is a selection technology that harnesses a massive oligonucleotide-peptide hybrid library to identify molecules that bind to protein targets; automating the mRNA display workflow is a route towards the rapid development of novel cancer protein binding agents. A major roadblock to the microfluidic automation of mRNA display is the nonspecific adhesion of the many required enzyme, peptide, and oligonucleotide reagents to the channel surfaces.

To minimize or eliminate this adhesion, we have explored a range of SLA resin formulations based on vinyl monomers with various functional groups. After examining the achievable resolution and mechanical properties of each formulation, we characterized peptide, oligonucleotide, and protein adhesion and determined the degree to which adhered enzymes retained enzymatic activity. Low-adhesion SLA-printed modules were assembled to construct an automated system capable of producing new mRNA display binding ligands.