Paper-based loop-mediated isothermal amplification and CRISPR integrated platform for on-site nucleic acid testing of pathogens.

We report the development and initial validation of a paper-based nucleic acid testing platform that integrates Loop-mediated isothermal amplification (LAMP) with clustered regularly interspaced short palindromic repeats (CRISPR) technology, referred to as PLACID (Paper-based LAMP-CRISPR Integrated Diagnostics). LAMP eliminates the need for thermal cycling, resulting in simplified instrumentation, and the CRISPR-associated protein (Cas 12a) system eliminates false positive signals from LAMP products, resulting in highly selective and sensitive assays. We optimized the assay to perform both amplification and detection entirely on paper, eliminating the need for complex fluid handling steps and lateral flow assay transfers.

Enhancing the Sensitivity of Lateral Flow Assay with Europium Nanoparticles for Accurate Human IgG Quantification.

Accurate quantification of immunoglobulin G (IgG) levels is vital for understanding immune status and diagnosing various medical conditions. Lateral flow assays (LFAs) offer rapid and convenient diagnostic tools, but their sensitivity has been a limitation. Our research introduces a refined method incorporating europium nanoparticles, enhancing both sensitivity and accuracy of LFAs in human IgG measurement.

A Smartphone-Enabled Continuous Flow Digital Droplet LAMP Platform for High Throughput and Inexpensive Quantitative Detection of Nucleic Acid Targets.

Molecular tests for infectious diseases and genetic anomalies, which account for significant global morbidity and mortality, are central to nucleic acid analysis. In this study, we present a digital droplet LAMP (ddLAMP) platform that offers a cost-effective and portable solution for such assays. Our approach integrates disposable 3D-printed droplet generator chips with a consumer smartphone equipped with a custom image analysis application for conducting ddLAMP assays, thereby eliminating the necessity for expensive and complicated photolithographic techniques, optical microscopes, or flow cytometers.

Miniaturized Non-Contact Heating and Transmitted Light Imaging Using an Inexpensive and Modular 3D-Printed Platform for Molecular Diagnostics.

The ability to simultaneously heat and image samples using transmitted light is crucial for several biological applications. However, existing techniques such as heated stage microscopes, thermal cyclers equipped with imaging capabilities, or non-contact heating systems are often bulky, expensive, and complex. This work presents the development and characterization of a Miniaturized Optically-clear Thermal Enclosure (MOTE) system-an open-source, inexpensive, and low-powered modular system-capable of convectively heating samples while simultaneously imaging them with transmitted light.

Carbon dots for photothermal applications.

Carbon dots are zero-dimensional nanomaterials that have garnered significant research interest due to their distinct optical properties, biocompatibility, low fabrication cost, and eco-friendliness. Recently, their light-to-heat conversion ability has led to several novel photothermal applications. In this minireview, we categorize and describe the photothermal application of carbon dots along with methods incorporated to enhance their photothermal efficiency.

Multi-Resin Masked Stereolithography (MSLA) 3D Printing for Rapid and Inexpensive Prototyping of Microfluidic Chips with Integrated Functional Components.

Stereolithography based 3D printing of microfluidics for prototyping has gained a lot of attention due to several advantages such as fast production, cost-effectiveness, and versatility over traditional photolithography-based microfabrication techniques. However, existing consumer focused SLA 3D printers struggle to fabricate functional microfluidic devices due to several challenges associated with micron-scale 3D printing. Here, we explore the origins and mechanism of the associated failure modes followed by presenting guidelines to overcome these challenges.

Effects of Relative Humidity and Paper Geometry on the Imbibition Dynamics and Reactions in Lateral Flow Assays.

Lateral flow assays and paper microfluidics have the potential to replace benchtop instrumented medical diagnostic systems with instrument-free systems that rely on passive transport of liquid through micro-porous paper substrates. Predicting the imbibition dynamics of liquid through dry paper substrates is mostly modeled through the Lucas-Washburn (LW) equations. However, the LW framework assumes that the fluid front exhibits a sharp boundary between the dry and wet phases across the liquid imbibition interface.

Synergistic Effect of Nitrogen Doping and Ultra-Microporosity on the Performance of Biomass and Microalgae-Derived Activated Carbons for CO Capture.

We report a unique naturally derived activated carbon with optimally incorporated nitrogen functional groups and ultra-microporous structure to enable high CO adsorption capacity. The coprocessing of biomass ( waste leaves) and microalgae (Spirulina) as the N-doping agent was investigated by probing the parameter space (biomass/microalgae weight ratio, reaction temperature, and reaction time) of hydrothermal carbonization and activation process (via the ZnCl/CO activation) to generate hydrochars and activated carbons, respectively, with tunable nitrogen content and pore sizes. The central composite-based design of the experiment was applied to optimize the parameters of the prehydrothermal carbonization procedure resulting in the fabrication of N-enriched carbonaceous products with the highest possible mass yield and nitrogen content.

Colorimetric-Luminance Readout for Quantitative Analysis of Fluorescence Signals with a Smartphone CMOS Sensor.

Smartphones have shown promise as an enabling technology for portable and distributed point-of-care diagnostic tests. The CMOS camera sensor can be used for detecting optical signals, including fluorescence for applications such as isothermal nucleic acid amplification tests. However, such analysis is typically limited mostly to end point detection of single targets.

Impact of primer dimers and self-amplifying hairpins on reverse transcription loop-mediated isothermal amplification detection of viral RNA.

Loop-mediated isothermal amplification (LAMP), coupled with reverse transcription (RT), has become a popular technique for detection of viral RNA due to several desirable characteristics for use in point-of-care or low-resource settings. The large number of primers in LAMP (six per target) leads to an increased likelihood of primer dimer interactions, and the inner primers in particular are prone to formation of stable hairpin structures due to their length (typically 40-45 bases). Although primer dimers and hairpin structures are known features to avoid in nucleic acid amplification techniques, there is little quantitative information in literature regarding the impact of these structures on LAMP or RT-LAMP assays.

A smartphone-based diagnostic platform for rapid detection of Zika, chikungunya, and dengue viruses.

Current multiplexed diagnostics for Zika, dengue, and chikungunya viruses are situated outside the intersection of affordability, high performance, and suitability for use at the point-of-care in resource-limited settings. Consequently, insufficient diagnostic capabilities are a key limitation facing current Zika outbreak management strategies. Here we demonstrate highly sensitive and specific detection of Zika, chikungunya, and dengue viruses by coupling reverse-transcription loop-mediated isothermal amplification (RT-LAMP) with our recently developed quenching of unincorporated amplification signal reporters (QUASR) technique.

Smartphone-Enabled Detection Strategies for Portable PCR-Based Diagnostics.

Incredible progress continues to be made toward development of low-cost nucleic acid-based diagnostic solutions suitable for deployment in resource-limited settings. Detection components play a vitally important role in these systems, but have proven challenging to adapt for operation in a portable format. Here we describe efforts aimed at leveraging the capabilities of consumer-class smartphones as a convenient platform to enable detection of nucleic acid products associated with DNA amplification via the polymerase chain reaction (PCR).

Synchronized chaotic targeting and acceleration of surface chemistry in prebiotic hydrothermal microenvironments.

Porous mineral formations near subsea alkaline hydrothermal vents embed microenvironments that make them potential hot spots for prebiotic biochemistry. But, synthesis of long-chain macromolecules needed to support higher-order functions in living systems (e.g.

A simple check valve for microfluidic point of care diagnostics.

Check valves are often essential components in microfluidic devices, enabling automated sample processing for diagnostics at the point of care. However, there is an unmet need for a check valve design that is compatible with rigid thermoplastic devices during all stages of development-from initial prototyping with a laser cutter to final production with injection molding. Here, we present simple designs for a passive, normally closed check valve that is manufactured from commonly available materials with a CO laser and readily integrated into prototype and production thermoplastic devices.

Lab-on-a-Drone: Toward Pinpoint Deployment of Smartphone-Enabled Nucleic Acid-Based Diagnostics for Mobile Health Care.

We introduce a portable biochemical analysis platform for rapid field deployment of nucleic acid-based diagnostics using consumer-class quadcopter drones. This approach exploits the ability to isothermally perform the polymerase chain reaction (PCR) with a single heater, enabling the system to be operated using standard 5 V USB sources that power mobile devices (via battery, solar, or hand crank action). Time-resolved fluorescence detection and quantification is achieved using a smartphone camera and integrated image analysis app.

Microscale chaotic advection enables robust convective DNA replication.

The ability of chaotic advection under microscale confinement to direct chemical processes along accelerated kinetic pathways has been recognized for some time. However, practical applications have been slow to emerge because optimal results are often counterintuitively achieved in flows that appear to possess undesirably high disorder. Here we present a 3D time-resolved analysis of polymerase chain reaction (PCR)-mediated DNA replication across a broad ensemble of geometric states.

Education: DNA replication using microscale natural convection.

There is a need for innovative educational experiences that unify and reinforce fundamental principles at the interface between the physical, chemical, and life sciences. These experiences empower and excite students by helping them recognize how interdisciplinary knowledge can be applied to develop new products and technologies that benefit society. Microfluidics offers an incredibly versatile tool to address this need.