Microfluidic Integration of Magnetically Functionalized Microwires for Flow Cytometry Protein Quantification.

A novel approach to protein quantification utilizing a microfluidic platform activated by a magnetic assembly of functionalized magnetic beads around soft magnetic capture centers is presented. Functionalized magnetic beads, known for their high surface area and facile manipulation under external magnetic fields, are injected inside microfluidic channels and immobilized magnetically on the surface of glass-coated soft magnetic microwires placed along the symmetry axis of these channels. A fluorescent (Cy5) immunomagnetic sandwich ELISA is then performed by sequentially flowing the sample and all necessary reagents in the microfluidic channels.

Combinatorial Nanoparticle-Bound ssDNA Oligonucleotide Library Synthesized by Split-and-Pool Synthesis.

Synthetic ssDNA oligonucleotides hold great potential for various applications, including DNA aptamers, DNA digital data storage, DNA origami, and synthetic genomes. In these contexts, precise control over the synthesis of the ssDNA strands is essential for generating combinatorial sequences with user-defined parameters. Desired features for creating synthetic DNA oligonucleotides include easy manipulation of DNA strands, effective detection of unique DNA sequences, and a straightforward mechanism for strand elongation and termination.

Microfluidic methods for the diagnosis of acute respiratory tract infections.

Acute respiratory tract infections (ARTIs) are caused by sporadic or pandemic outbreaks of viral or bacterial pathogens, and continue to be a considerable socioeconomic burden for both developing and industrialized countries alike. Diagnostic methods and technologies serving as the cornerstone for disease management, epidemiological tracking, and public health interventions are evolving continuously to keep up with the demand for higher sensitivity, specificity and analytical throughput. Microfluidics is becoming a key technology in these developments as it allows for integrating, miniaturizing and automating bioanalytical assays at an unprecedented scale, reducing sample and reagent consumption and improving diagnostic performance in terms of sensitivity, throughput and response time.

Sample-to-answer centrifugal microfluidic droplet PCR platform for quantitation of viral load.

Droplet digital polymerase chain reaction (ddPCR) stands out as a highly sensitive diagnostic technique that is gaining traction in infectious disease diagnostics due to its ability to quantitate very low numbers of viral gene copies. By partitioning the sample into thousands of droplets, ddPCR enables precise and absolute quantification without relying on a standard curve. However, current ddPCR systems often exhibit relatively low levels of integration, and the analytical process remains dependent on elaborate workflows for up-front sample preparation.

Reversible bonding in thermoplastic elastomer microfluidic platforms for harvestable 3D microvessel networks.

Transplantable ready-made microvessels have therapeutic potential for tissue regeneration and cell replacement therapy. Inspired by the natural rapid angiogenic sprouting of microvessels , engineered injectable 3D microvessel networks are created using thermoplastic elastomer (TPE) microfluidic devices. The TPE material used here is flexible, optically transparent, and can be robustly yet reversibly bonded to a variety of plastic substrates, making it a versatile choice for microfluidic device fabrication because it overcomes the weak self-adhesion properties and limited manufacturing options of poly(dimethylsiloxane) (PDMS).

Centrifugal microfluidic system for colorimetric sample-to-answer detection of viral pathogens.

We describe a microfluidic system for conducting thermal lysis, polymerase chain reaction (PCR) amplification, hybridization, and colorimetric detection of foodborne viral organisms in a sample-to-answer format. The on-chip protocol entails 24 steps which are conducted by a centrifugal platform that allows for actuating liquids pneumatically during rotation and so facilitates automation of the workflow. The microfluidic cartridge is fabricated from transparent thermoplastic polymers and accommodates assay components along with an embedded micropillar array for detection and read-out.

Automated sample-to-answer centrifugal microfluidic system for rapid molecular diagnostics of SARS-CoV-2.

Testing for SARS-CoV-2 is one of the most important assets in COVID-19 management and mitigation. At the onset of the pandemic, SARS-CoV-2 testing was uniquely performed in central laboratories using RT-qPCR. RT-qPCR relies on trained personnel operating complex instrumentation, while time-to-result can be lengthy (, 24 to 72 h).

Use of Polymer Micropillar Arrays as Templates for Solid-Phase Immunoassays.

We investigate the use of periodic micropillar arrays produced by high-fidelity microfabrication with cyclic olefin polymers for solid-phase immunoassays. These three-dimensional (3D) templates offer higher surface-to-volume ratios than two-dimensional substrates, making it possible to attach more antibodies and so increase the signal obtained by the assay. Micropillar arrays also provide the capacity to induce wicking, which is used to distribute and confine antibodies on the surface with spatial control.

On-the-Fly Phase Transition and Density Changes of Aqueous Two-Phase Systems on a Centrifugal Microfluidic Platform.

This paper describes on-the-fly physical property changes of aqueous two-phase systems (ATPS) in microfluidic devices. The properties and phases of the ATPS are modulated on-demand by using a centrifugal microfluidic device filled with poly(ethylene glycol) (PEG) and dextran (DEX) solutions. By use of the centrifugal force and active pneumatic controls provided by a centrifugal microfluidic platform (CMP), PEG-DEX mixtures are manipulated and processed inside simple thermoplastic microfluidic devices.

Multifunctional magnetic nanoparticle cloud assemblies for capture of bacteria and isolation of microbial DNA.

We investigate the formation of suspended magnetic nanoparticle (MNP) assemblies (M-clouds) and their use for bacterial capture and DNA extraction. M-clouds are obtained as a result of magnetic field density variations when magnetizing an array of micropillars coated with a soft ferromagnetic NiP layer. Numerical simulations suggest that the gradient in the magnetic field created by the pillars is four orders of magnitude higher than the gradient generated by the external magnets.

Real-time monitoring of bead-based DNA hybridization in a microfluidic system: study of amplicon hybridization behavior on solid supports.

DNA hybridization phenomena occurring on solid supports are not understood as clearly as aqueous phase hybridizations and mathematical models cannot predict some empirically obtained results. Ongoing research has identified important parameters but remains incomplete to accurately account for all interactions. It has previously been shown that the length of the overhanging (dangling) end of the target DNA strand following hybridization to the capture probe is correlated to interactions with the complementary strand in solution which can result in unbinding of the target and its release from the surface.

Evaporation-Driven Water-in-Water Droplet Formation.

We present new observations of aqueous two-phase system (ATPS) thermodynamic and interfacial phenomena that occur inside sessile droplets due to water evaporation. Sessile droplets that contain polymeric solutions, which are initially in equilibrium in a single phase, are observed at their three-phase liquid-solid-air contact line. As evaporation of a sessile droplet proceeds, we find that submicron secondary water-in-water (W/W) droplets emerge spontaneously at the edges of the mother sessile droplet due to the resulting phase separation from water evaporation.

Centrifugal microfluidic lab-on-a-chip system with automated sample lysis, DNA amplification and microarray hybridization for identification of enterohemorrhagic culture isolates.

The development of technology for the rapid, automated identification of bacterial culture isolates can help regulatory agencies to shorten response times in food safety surveillance, compliance, and enforcement as well as outbreak investigations. While molecular methods such as polymerase chain reaction (PCR) enable the identification of microbial organisms with high sensitivity and specificity, they generally rely on sophisticated instrumentation and elaborate workflows for sample preparation with an undesirably high level of hands-on engagement. Herein, we describe the design, operation and performance of a lab-on-a-chip system integrating thermal lysis, PCR amplification and microarray hybridization on the same cartridge.

Methylation Specific Multiplex Droplet PCR using Polymer Droplet Generator Device for Hematological Diagnostics.

A multiplexed droplet PCR (mdPCR) workflow and detailed protocol for determining epigenetic-based white blood cell (WBC) differential count is described, along with a thermoplastic elastomer (TPE) microfluidic droplet generation device. Epigenetic markers are used for WBC subtyping which is of important prognostic value in different diseases. This is achieved through the quantification of DNA methylation patterns of specific CG-rich regions in the genome (CpG loci).

Buoyancy-driven step emulsification on pneumatic centrifugal microfluidic platforms.

We present here a new method for controlling the droplet size in step emulsification processes on a centrifugal microfluidic platform, which, in addition to the centrifugal force, uses pneumatic actuation for fluid displacement. We highlight the importance of the interplay between buoyancy effects and the flow rate at the step junction, and provide a simple analytical model relating these two quantities to the size of the droplets. Numerical models as well as experiments with water-in-oil emulsions are performed in support of the proposed model.

Polymer Micropillar Arrays for Colorimetric DNA Detection.

We describe the use of periodic micropillar arrays, produced from cyclic olefin copolymer using high-fidelity microfabrication, as templates for colorimetric DNA detection. The assay involves PCR-amplified gene markers for O157:H7 (, , , and ) incorporating a detectable digoxigenin label, which is revealed through an immunoenzymatic process following hybridization with target-specific oligonucleotide capture probes. The capacity of micropillar arrays to induce wicking is used to distribute and confine capture probes with spatial control, making it possible to achieve a uniform signal while allowing multiple, independent probes to be arranged in close proximity on the same substrate.

Epigenetic subtyping of white blood cells using a thermoplastic elastomer-based microfluidic emulsification device for multiplexed, methylation-specific digital droplet PCR.

Epigenetic markers attract increasing attention for the study of phenotypic variations, which has led to the investigation of cell-lineage DNA methylation patterns that correlate with human leukocyte populations for obtaining counts of white blood cell (WBC) subsets. Current methods of DNA methylation analysis involve genome sequencing or loci-specific quantitative PCR (qPCR). Herein, a multiplexed digital droplet PCR (ddPCR) workflow for determining epigenetic-based WBC differential count is described for the first time.

Antibody Response and Disease Severity in Healthcare Worker MERS Survivors.

We studied antibody response in 9 healthcare workers in Jeddah, Saudi Arabia, who survived Middle East respiratory syndrome, by using serial ELISA and indirect immunofluorescence assay testing. Among patients who had experienced severe pneumonia, antibody was detected for >18 months after infection. Antibody longevity was more variable in patients who had experienced milder disease.

Polymer-based microfluidic chip for rapid and efficient immunomagnetic capture and release of Listeria monocytogenes.

Infections caused by foodborne pathogens such as Listeria monocytogenes pose a threat to public health while timely detection is challenging due to pathogen low numbers. The development of robust and efficient sample preparation techniques is crucial to improve detection sensitivity and workflow. Immunomagnetic separation using magnetic nanoparticles (MNPs) is attractive, as it can efficiently capture target cells.

From cellular lysis to microarray detection, an integrated thermoplastic elastomer (TPE) point of care Lab on a Disc.

We present an all-thermoplastic integrated sample-to-answer centrifugal microfluidic Lab-on-Disc system (LoD) for nucleic acid analysis. The proposed CD system and engineered platform were employed for analysis of Bacillus atrophaeus subsp. globigii spores.

All-thermoplastic nanoplasmonic microfluidic device for transmission SPR biosensing.

Early and accurate disease diagnosis still remains a major challenge in clinical settings. Biomarkers could potentially provide useful tools for the detection and monitoring of disease progression, treatment safety and efficacy. Recent years have witnessed prodigious advancement in biosensor development with research directed towards rapid, real-time, label-free and sensitive biomarker detection.

Stigmatization of 'psychiatric label' by medical and non-medical students.

Background: Stigmatization of psychiatric patients is present both in the general population and among healthcare professionals.

Aim: To determine the attitudes and behaviour of medical students towards a person who goes to a psychiatrist, before and after psychiatric rotation, and to compare those attitudes between medical and non-medical students.

Methods: The study included 525 medical students (second and sixth year of studies) and 154 students of law.

Designed biointerface using near-infrared quantum dots for ultrasensitive surface plasmon resonance imaging biosensors.

The surface plasmon resonance imaging chip biointerface is fully designed using near-infrared (NIR) quantum dots (QDs) for the enhancement of surface plasmon resonance imaging (SPRi) signals in order to extend their application for medical diagnostics. The measured SPRi detection signal following the QD binding to the surface was amplified 25-fold for a 1 nM concentration of single-stranded DNA (ssDNA) and 50-fold for a 1 μg/mL concentration of prostate-specific antigen (PSA), a cancer biomarker, thus substantiating their wide potential to study interactions of a diverse set of small biomolecules. This significant enhancement is attributed to the QD's mass-loading effect and spontaneous emission coupling with propagating surface plasmons, which allowed the SPRi limit of detection to be reduced to 100 fM and 100 pg/mL for ssDNA and PSA, respectively.

Nanostructured digital microfluidics for enhanced surface plasmon resonance imaging.

The advances in genomics and proteomics have unveiled an exhaustive catalogue of biomarkers that can potentially be used as diagnostic and prognostic indicators of genetic and infectious diseases. Current thrust in biosensor development is towards rapid, real-time, label-free and highly sensitive detection of the indicative biomarkers. While surface plasmon resonance imaging (SPRi) biosensors could potentially be the best suited candidate for biomarker-based diagnosis, important milestones need to be reached.

Integration and detection of biochemical assays in digital microfluidic LOC devices.

The ambition of lab-on-a-chip (LOC) systems to achieve chip-level integration of a complete analytical process capable of performing a complex set of biomedical protocols is hindered by the absence of standard fluidic components able to be assembled. As a result, most microfluidic platforms built to date are highly specialized and designed to fulfill the requirements of a single particular application within a limited set of operations. Electrowetting-on-dielectric (EWOD) digital microfluidic technology has been recently introduced as a new methodology in the quest for LOC systems.