Innovations in Microfluidics & SCA 2021 Agenda

Conference Registration, Materials Pick-Up, Morning Coffee and Pastries

Diffusion-based Separation Using Bidirectional Electroosmotic Flow
Moran Bercovici, Associate Professor, Faculty of Mechanical Engineering; Head, Technion Microfluidic Technologies Laboratory, Technion, Israel Institute of Technology, Israel

We present a microscale separation method that leverages bidirectional flow, generated by an array of alternate-current field-effect electrodes, to electroosmotically tune the dispersion regime of molecules and particles. Under bidirectional flow the relative motion of species due to differences in their molecular diffusivity can be significantly enhanced. The system can be configured so that low diffusivity species experience a ballistic transport regime and are advected through the chamber, whereas high diffusivity species experience a diffusion dominated regime with zero average velocity and are retained in the chamber.  We experimentally demonstrate the separation of particles, antibodies, and dyes, and present a theoretical analysis of the system, providing engineering guidelines for its optimal design and operation. This method provides means for leveraging molecular diffusivity for analysis and sample preparation applications, particularly for sub-microliter sample volumes that are not compatible with standard separation techniques.

Morning Coffee Break and Networking

Customizing Single Cell Research
Thomas Ayers, Sr. Technical Applications Specialist, Dolomite Bio

Here we report a flexible and cost-effective droplet-based microfluidics system, called the Nadia Instrument, that can allow 3’ mRNA capture of ~50,000 single cells or individual nuclei in a single run. The precise pressure-based system demonstrates highly reproducible droplet size, low doublet rates and high mRNA capture efficiencies that compare favourably in the field. Moreover, when combined with the Nadia Innovate, the system can be transformed into an adaptable setup that enables use of different buffers and barcoded bead configurations to facilitate diverse applications. We demonstrate how this robust and versatile system unlocks high throughput single cell profiling for large plant leaf protoplasts and single cells or nuclei with poor RNA yields. With the Nadia Innovate, we present a solution for diverse sample types, whilst allowing researchers to complete more single cell transcriptomic projects within budget.

Networking Lunch in the Exhibit Hall for the Physical On-Site Participants

YAMAHA CELL HANDLER: Automated, Rapid Spheroid, Organoid and Single Cell Sort System by Imaging and Picking
Yuichi Hikichi, General Manager, MDB Division , Yamaha Motor

By using various parameters of images, like size, shape and fluorescent intensity, the CELL HANDLER^TM can automatically select single cells or 3D spheroids and sort them to various types of culture plates. It uses a unique disposable tip to ensure the damage-free and accurate handling and enables direct picking of organoids from gel. Conventional manual methods have been hampered by low throughput and suboptimal accuracy. Conventional cell sorters damage cells and need significant amounts of cells and are not applicable for spheroids and organoids. The CELL HANDLER complements these deficiencies. It enables more accurate and higher throughput sorting than manual operation and low amounts of cells (e.g. 100-1000 cells) can be sorted without damages. Any types of plates (e.g. microtiter plates, micro cavity plates, 3D colony in gel, 2D colony on plate, organ-on-a-chip) can be used as a source plate and destination plate, which will support many types of applications. The CELL HANDLER is a high-speed, high-precision pick and place system coupled with image capture and analysis capability that greatly increases the efficiency and reliability of spheroid/organoid and single cell assays.

A Systematic Comparison of Single Cell RNA-Seq Methods
Joshua Levin, Senior Group Leader, Research Scientist, Stanley Center for Psychiatric Research, Klarman Cell Observatory, The Broad Institute of MIT and Harvard, United States of America

A multitude of single-cell RNA sequencing methods have been developed in recent years, with dramatic advances in scale and power, and enabling major discoveries and large scale cell mapping efforts. We directly compared seven methods for single cell and/or single nucleus profiling from three types of samples – cell lines, peripheral blood mononuclear cells and brain tissue. To analyze these datasets, we developed and applied scumi, a flexible computational pipeline that can be used for any scRNA-seq method. We evaluated the methods for both basic performance and for their ability to recover known biological information in the samples.

Single-Button Nucleic Acid Extraction for Rapid, Reliable Molecular Testing
Richard Chasen Spero, CEO, Redbud Labs

There are myriad exciting microfluidic platforms for molecular analysis, but few are designed to work with raw samples. We describe a new initiative at Redbud Labs to enable single-button sample preparation on a microfluidic cartridge, focusing on nucleic acid extraction for amplification and sequencing applications. Our method combines gold-standard laboratory biochemistry with cartridge-ready™ chips, MXR™DryPak and STR™BeadPak, to deliver lab-quality yield, fast runtimes, and compact design. We will also share strategies employed to develop our platform on a highly compressed timeline.

Afternoon Break

Microfluidics’ New Wave: Digitally-Fabricated Microdevices
Albert Folch, Professor of Bioengineering, University of Washington, United States of America

The vast majority of microfluidic devices are presently manufactured using micromolding processes that work very well for a reduced set of biocompatible materials, but the time, cost, and design constraints of micromolding hinder the commercialization of many devices. PDMS, in particular, is extremely popular in academic labs, yet the fabrication procedures are based on cumbersome manual methods and the material itself strongly absorbs lipophilic drugs. As a result, the dissemination of many cell-based microfluidic chips – and their impact on society – is in jeopardy. Digital Manufacturing (DM) is a family of computer-centered processes that integrate digital 3D designs, automated (additive or subtractive) fabrication, and device testing in order to increase fabrication efficiency. Importantly, DM enables the inexpensive realization of 3D designs that are impossible or very difficult to mold. The adoption of DM by microfluidic engineers has been slow, likely due to concerns over the resolution of the printers and the biocompatibility of the resins. We have developed microfluidic devices by SL in PEG-DA-based resins with automation and biocompatibility ratings similar to those made with PDMS. The resins allow for building transparent microchannels, microvalves, and multi-material devices containing hydrogels of larger-MW PEG-DA formulations.

The Simultaneous Determination of Biomolecules Binding Affinities On Multiple Varied Immobilization Surfaces
Claude Dufresne, CEO, axiVEND LLC

In many microfluidics devices, biomolecules are captured by pre-deposited proteins in channels, reaction chambers, or on microarrays. The correct immobilization of these proteins is critical to obtain the best device performance.  While many polymeric materials exist for protein immobilization, it is often quite tedious to go through a selection process.  Beyond binding capacity as a classic factor, often the orientation and functional ligand activity are more important.

This talk will present a method by which multiple polymers can be evaluated in parallel, in the same experiment.  Dozens of candidate surface binding polymeric materials are spotted on a Si/SiO2 chip, followed by spot-on-spot protein functionalization.  Target protein capture is then observed by interferometric reflectance. This allows for binding curves to be generated for each protein/surface polymer pair, in parallel, and under the same conditions.  The results are invaluable in assessing various polymeric formulations for optimal presentation of the capture molecule and its ensuing target molecule binding properties.

Since the underlying device material is covered with the binding polymer, the method described is applicable to a wide range of applications.

Highly Multiplexed Diagnostics with Droplet Microfluidics Enhanced by Compressed Sensing
Pavan Kota, Ph.D. Student, Dept of Bioengineering, Rice University, United States of America

Microfluidics can generate thousands of droplets to capture individual analytes, but usually only a few measurements can be acquired from each droplet at high throughput. Multiplexing efficiently with limited sensors is possible through compressed sensing if samples are sparse; most possible analytes must be absent from any particular sample. The authors recently developed a new compressed sensing algorithm called Sparse Poisson Recovery (SPoRe) that further exploits the Poisson statistics of microfluidic capture. Given an application-driven measurement model, SPoRe efficiently solves a maximum likelihood estimation problem to recover total analyte abundances. This work presents the first in vitro demonstration of SPoRe towards bacterial infection diagnostics with 16S droplet digital PCR (ddPCR). Five nonspecific probes assign binary barcodes to nine bacterial genera. Each droplet measurement is modeled as an OR operation among the present 16S genes’ barcodes. Although a single droplet’s contents may be ambiguous under this model, SPoRe solves for bacterial abundances by considering all droplets simultaneously. Moreover, SPoRe can pool data from multiple reactions with different subsets of probes to address limitations caused by probe cross-reactivity and fluorescence spectral overlap. SPoRe raises new possibilities in ddPCR-based diagnostics, and its modularity for nearly any measurement model enables applications beyond digital sensing.

Rapid Pre-Concentration for Salivary Measurement of Respiratory Viruses such as Influenza and COVID
Amy Drexelius, PhD Candidate, University of Cincinnati, United States of America

If there were a simple and passive technique, that would downshift the range of detection of all your sensors and analytes by 10-100X, what value would you place on such a breakthrough? Presented are single step and fast membrane pre-concentration devices for applications ranging from continuous wearable biosensing (sweat, interstitial fluid) to point of care diagnostics (blood, urine, saliva), which can detect analytes present in respiratory viruses such as influenza and COVID-19.  These devices now show >10X preconcentration in minutes for analytes ranging from small molecules (e.g. cortisol) to large proteins (influenza).  Further demonstrated are techniques that allow rapid adaptation of the devices for a wide variety of sensing modalities, including those sensitive to changes in pH or salinity.

Characterization and Modeling of Paper-based Bi-Material Actuator Cantilever; Application in Phosphate Detection
Amer Charbaji, Researcher, University of Rhode Island, United States of America

Paper-based microfluidics provide a simple, inexpensive, and user friendly approach for a variety of applications, such as environmental monitoring, healthcare, food safety, and various other miscellaneous purposes. Paper-based devices are made up of several different sections and, depending on the application, may require the use of an actuator for proper control of flow time, reaction time, sample volume or the sequential loading of reagents. In our previous publication, we described a novel autonomously-activated bi-material actuator cantilever (B-MAC) that offers several unique advantages for use in paper-based microfluidic devices. The B-MAC allows for the autonomous operation of paper-based devices that require sequential and timed delivery of multiple reagents. It is also simple to fabricate, requires small sample volumes to operate, and has a quick response time. The B-MAC is made up of a strip of filter paper that is partially laminated on one side with tape. Once the filter paper side is exposed to a fluid, the cellulose fibers experience hygro-expansion, whereas the adjoining tape layer remains constant in length. This difference in the expansion behavior of the paper and tape cause the cantilever to bend in the direction of the tape layer as shown below and touch the stationary component (SC). The B-MAC has been shown to be applicable to 1D, 2D, and 3D lateral flow devices. In this study, the B-MAC will be assessed as a potential actuator to be used in a paper-based device for the sensitive detection of phosphate in water. The most commonly used method to determine soluble phosphate in water is based on the molybdenum blue reaction. This assay has been implemented on paper-based devices before and requires multi-step loading of reagents to improve sensitivity, shelf life, and to reduce auto-reduction of the reagents. Using B-MAC in a paper-based device for phosphate detection will allow the sequential loading of multiple reagents to perform the molybdenum blue assay to detect phosphate with minimal human intervention other than simply adding the sample fluid.

Close of Day 1 of the Conference