Parallelization of Curved Inertial Microfluidic Channels to Increase the Throughput of Simultaneous Microparticle Separation and Washing.

The rising global need for clean water highlights the importance of efficient sample preparation methods to separate and wash various contaminants such as microparticles. Microfluidic methods for these purposes have emerged but they mostly deliver either separation or washing, with very low throughputs. Here, we investigate parallelization of a curved-channel particle separation and washing device in order to increase its throughput for sample preparation. A curved microchannel applies inertial forces to focus larger 10 µm microparticles at the inner wall of the channel and separate them from smaller 5 µm microparticles at the outer wall. At the same time, Dean flow recirculation is used to exchange the carrier solution of the large microparticles to a clean buffer (washing). We increased the number of curved channels in a stepwise manner from two to four to eight channels in two different arraying designs, i.e., rectangular and polar arrays. We examined efficient separation of target 10 µm particles from 5 µm particles, while transferring the larger microparticles into a clean buffer. Dean flow recirculation studies demonstrated that the rectangular arrayed device performs better, providing solution exchange efficiencies of more than 96% on average as compared to 89% for the polar array device. Our 8-curve rectangular array device provided a particle separation efficiency of 98.93 ± 0.91%, while maintaining a sample purity of 92.83 ± 1.47% at a high working flow rate of 12.8 mL/min. Moreover, the target particles were transferred into a clean buffer with a solution exchange efficiency of 96.81 ± 0.54% in our 8-curve device. Compared to the literature, our in-plane parallelization design of curved microchannels resulted in a 13-fold increase in the working flow rate of the setup while maintaining a very high performance in particle separation and washing. Our microfluidic device offers the potential to enhance the throughput and the separation and washing efficiencies in applications for biological and environmental samples.

Electrochemical Impedance Spectroscopy-Based Microfluidic Biosensor Using Cell-Imprinted Polymers for Bacteria Detection.

The rapid and sensitive detection of bacterial contaminants using low-cost and portable point-of-need (PoN) biosensors has gained significant interest in water quality monitoring. Cell-imprinted polymers (CIPs) are emerging as effective and inexpensive materials for bacterial detection as they provide specific binding sites designed to capture whole bacterial cells, especially when integrated into PoN microfluidic devices. However, improving the sensitivity and detection limits of these sensors remains challenging. In this study, we integrated CIP-functionalized stainless steel microwires (CIP-MWs) into a microfluidic device for the impedimetric detection of E. coli bacteria. The sensor featured two parallel microchannels with three-electrode configurations that allowed simultaneous control and electrochemical impedance spectroscopy (EIS) measurements. A CIP-MW and a non-imprinted polymer (NIP)-MW suspended perpendicular to the microchannels served as the working electrodes in the test and control channels, respectively. Electrochemical spectra were fitted with equivalent electrical circuits, and the charge transfer resistances of both cells were measured before and after incubation with target bacteria. The charge transfer resistance of the CIP-MWs after 30 min of incubation with bacteria was increased. By normalizing the change in charge transfer resistance and analyzing the dose-response curve for bacterial concentrations ranging from 0 to 107 CFU/mL, we determined the limits of detection and quantification as 2 × 102 CFU/mL and 1.4 × 104 CFU/mL, respectively. The sensor demonstrated a dynamic range of 102 to 107 CFU/mL, where bacterial counts were statistically distinguishable. The proposed sensor offers a sensitive, cost-effective, durable, and rapid solution for on-site identification of waterborne pathogens.

Simultaneous high-throughput particle-bacteria separation and solution exchange via in-plane and out-of-plane parallelization of microfluidic centrifuges.

Inertial microfluidic devices have gained attention for point-of-need (PoN) sample preparation. Yet, devices capable of simultaneous particle-bacteria solution exchange and separation are low in throughput, hindering their applicability to PoN settings. This paper introduces a microfluidic centrifuge for high-throughput solution exchange and separation of microparticles, addressing the need for processing large sample volumes at elevated flow rates. The device integrates Dean flow recirculation and inertial focusing of microparticles within 24 curved microchannels assembled in a three-layer configuration via in-plane and out-of-plane parallelization. We studied solution exchange and particle migration using singleplex and duplex samples across devices with varying curve numbers (2-curve, 8-curve, and 24-curve). Processing 5 and 10 µm microparticles at flow rates up to 16.8 ml/min achieved a solution exchange efficiency of 96.69%. In singleplex solutions, 10 and 5 µm particles selectively migrated to inner and outer outlets, demonstrating separation efficiencies of 99.7% and 90.3%, respectively. With duplex samples, sample purity was measured to be 93.4% and 98.6% for 10 and 5 µm particles collected from the inner and the outer outlets, respectively. Application of our device in biological assays was shown by performing duplex experiments where 10 µm particles were isolated from Salmonella bacterial suspension with purity of 97.8% while increasing the state-of-the-art particle solution exchange and separation throughput by 16 folds. This parallelization enabled desirable combinations of high throughput, low-cost, and scalability, without compromising efficiency and purity, paving the way for sample preparation at the PoN in the future.

Fabrication of Porous Collagen Scaffolds Containing Embedded Channels with Collagen Membrane Linings.

Tissues and organs contain an extracellular matrix (ECM). In the case of blood vessels, endothelium cells are anchored to a specialized basement membrane (BM) embedded inside the interstitial matrix (IM). We introduce a multi-structural collagen-based scaffold with embedded microchannels that mimics in vivo structures within vessels. Our scaffold consists of two parts, each containing two collagen layers, i.e., a 3D porous collagen layer analogous to IM lined with a thin 2D collagen film resembling the BM. Enclosed microchannels were fabricated using contact microprinting. Microchannel test structures with different sizes ranging from 300 to 800 µm were examined for their fabrication reproducibility. The heights and perimeters of the fabricated microchannels were ~20% less than their corresponding values in the replication PDMS mold; however, microchannel widths were significantly closer to their replica dimensions. The stiffness, permeability, and pore size properties of the 2D and 3D collagen layers were measured. The permeability of the 2D collagen film was negligible, making it suitable for mimicking the BM of large blood vessels. A leakage test at various volumetric flow rates applied to the microchannels showed no discharge, thereby verifying the reliability of the proposed integrated 2D/3D collagen parts and the contact printing method used for bonding them in the scaffold. In the future, multi-cell culturing will be performed within the 3D porous collagen and against the 2D membrane inside the microchannel, hence preparing this scaffold for studying a variety of blood vessel-tissue interfaces. Also, thicker collagen scaffold tissues will be fabricated by stacking several layers of the proposed scaffold.

Innovating Ferro-sonication approach for extracting microplastics from wastewater.

For accurate and reliable analysis of microplastics (MPs) in wastewater (WW), it is imperative to comprehend the significance of pre-treating WW before analysis. The suspended solids (SS) in the matrix tend to adhere to the MPs during filtration, which interferes with the detection of the MPs. In this regard, the present study aims to develop and optimize a pretreatment method to improve the extraction efficiency of MPs from WW by reducing the SS. A combination of the Fenton reaction and ultrasonication, ferro-sonication (Fe-UlS), was proposed to digest and eliminate the SS from WW. This hybrid pretreatment, Fe-UlS, was optimized for ultrasonication amplitude, treatment time, and hydrogen peroxide dose using response surface methodology (RSM) with a Box-Behnken design, achieving a desirability of 0.984. The optimum conditions for the Fe-UlS, such as the (1:1) Fenton reagent ratio (0.05 M FeSO4: 30 % H2O2), ultrasonication amplitude (31 %), and total process time (30 min) were found to be statistically significant (p < 0.05). The developed method was then employed for the extraction of spiked polyethylene (PE), polypropylene (PP) and polyethylene terephthalate (PET) MPs in real WW and found efficient in removing 83 % of the TSS present in the primary influent were in 30 min at a temperature of 45 °C. Also, the method did not affect the physio-chemical characteristics of the MPs; however, the thermal analysis of PE and PP MPs showed a statistically significant decrease in the melting temperature, as proven by paired t-test analysis. Further, a non-targeted liquid chromatography-mass spectrometry (LC-MS) analysis proved that Fe-UlS is a stable process, as it did not cause any leaching of MPs under the optimum pretreatment conditions. Finally, Laser Direct-Infrared Imaging (LD-IR) analysis was conducted to validate the developed Fe-UlS pretreatment approach for MP analysis in real WW. About 3434 MPs were detected in 100 mL of WW primary influent, within the size range of 9 to 500 µm. This hybrid pretreatment approach not only streamlines WW sample processing but also reduces the required concentration of Fenton reagent and processing time, yielding accurate and reliable results for monitoring MPs in WW.

High-throughput light sheet imaging of adult and larval C. elegans Parkinson's disease model using a low-cost optofluidic device and a fluorescent microscope.

Recent advancements at the interface of microfluidics technology and light sheet fluorescence microscopy have opened the door for high-throughput and high-content investigation of C. elegans disease models. In this paper, we report on the development of a simple, miniaturized, and low-cost optofluidic platform that can be added to a conventional inverted fluorescent microscope for continuous light sheet imaging of transgenic worm populations with high lateral and axial resolutions of 1.1 µm and 2.4 µm, respectively. The optofluidic device is made entirely of PDMS with integrated optics for light sheet generation. Laser excitation is delivered to the device via a low-cost free space laser, and cross-sections of worm populations are imaged as they pass continuously through a channel. Results show the platform can image NW1229 whole worms with pan-neural fluorescent expression at a throughput of >20 worms per minute at L3 and young adult (YA) stages. As a benchmark test, we show that the low-cost device can quantify the reduced neuronal expressions of L3 and YA NW1229 worms when exposed to 500 µM 6-OHDA neurodegenerative agent. Following the benchmark validation, we utilized the platform in a novel application for imaging human alpha-synuclein reporter in populations of Parkinson's transgenic model (ERS100). Results show the ability of the low-cost platform to reliably detect and quantify the anomalous neural phenotypic changes in ERS100 populations at L3 and YA stages with high spatial resolution. The findings of this study show the potential of our low-cost optofluidic add-on platform to equip conventional fluorescent microscopes with light sheet capability for quantitative phenotypic studies of transgenic C. elegans at high resolution and throughput.

Microfluidic Sensor Based on Cell-Imprinted Polymer-Coated Microwires for Conductometric Detection of Bacteria in Water.

The rapid, inexpensive, and on-site detection of bacterial contaminants using highly sensitive and specific microfluidic sensors is attracting substantial attention in water quality monitoring applications. Cell-imprinted polymers (CIPs) have emerged as robust, cost-effective, and versatile recognition materials with selective binding sites for capturing whole bacteria. However, electrochemical transduction of the binding event to a measurable signal within a microfluidic device to develop easy-to-use, compact, portable, durable, and affordable sensors remains a challenge. For this paper, we employed CIP-functionalized microwires (CIP-MWs) with an affinity towards E. coli and integrated them into a low-cost microfluidic sensor to measure the conductometric transduction of CIP-bacteria binding events. The sensor comprised two CIP-MWs suspended perpendicularly to a PDMS microchannel. The inter-wire electrical resistance of the microchannel was measured before, during, and after exposure of CIP-MWs to bacteria. A decline in the inter-wire resistance of the sensor after 30 min of incubation with bacteria was detected. Resistance change normalization and the subsequent analysis of the sensor's dose-response curve between 0 to 109 CFU/mL bacteria revealed the limits of detection and quantification of 2.1 × 105 CFU/mL and 7.3 × 105 CFU/mL, respectively. The dynamic range of the sensor was 104 to 107 CFU/mL where the bacteria counts were statistically distinguishable from each other. A linear fit in this range resulted in a sensitivity of 7.35 µS per CFU/mL. Experiments using competing Sarcina or Listeria cells showed specificity of the sensor towards the imprinted E. coli cells. The reported CIP-MW-based conductometric microfluidic sensor can provide a cost-effective, durable, portable, and real-time solution for the detection of pathogens in water.

Loss of Panx1 function in zebrafish alters motor behavior in a lab-on-chip model of Parkinson's disease.

Pannexin 1 (Panx1) forms ATP-permeable membrane channels that play roles in purinergic signaling in the nervous system. A link between Panx1 activity and neurodegenerative disorders including Parkinson's disease (PD) has been suggested, but experimental evidence is limited. Here, a zebrafish model of PD was produced by exposing panx1a+/+ and panx1a-/- zebrafish larvae to 6-hydroxydopamine (6-OHDA). Electrical stimulation in a microfluidic chip and quantitative real-time-qPCR of zebrafish larvae tested the role of Panx1 in both pathological and normal conditions. After 72-h treatment with 6-OHDA, the electric-induced locomotor activity of 5 days post fertilization (5dpf) panx1a+/+ larvae were reduced, while the stimulus did not affect locomotor activity of age-matched panx1a-/- larvae. A RT-qPCR analysis showed an increase in the expression of genes that are functionally related to dopaminergic signaling, like the tyrosine hydroxylase (th2) and the leucine-rich repeat kinase 2 (lrrk2). Extending the 6-OHDA treatment duration to 120 h caused a significant reduction in the locomotor response of 7dpf panx1a-/- larvae compared to the untreated panx1a-/- group. The RT-qPCR data showed a reduced expression of dopaminergic signaling genes in both genotypes. It was concluded that the absence of Panx1a channels compromised dopaminergic signaling in 6-OHDA-treated zebrafish larvae and that the increase in the expression of dopaminergic genes was transient, most likely due to a compensatory upregulation. We propose that zebrafish Panx1a models offer opportunities to shed light on PD's physiological and molecular basis. Panx1a might play a role on the progression of PD, and therefore deserves further investigation.

Recent advances in biosensors for real time monitoring of pH, temperature, and oxygen in chronic wounds.

Chronic wounds are among the major healthcare issues affecting millions of people worldwide with high rates of morbidity, losses of limbs and mortality. Microbial infection in wounds is a severe problem that can impede healing of chronic wounds. Accurate, timely and early detection of infections, and real time monitoring of various wound healing biomarkers related to infection can be significantly helpful in the treatment and care of chronic wounds. However, clinical methodologies of periodic assessment and care of wounds require physical visit to wound care clinics or hospitals and time-consuming frequent replacement of wound dressing patches, which also often adversely affect the healing process. Besides, frequent replacements of wound dressings are highly expensive, causing a huge amount of burden on the national health care systems. Smart bandages have emerged to provide in situ physiochemical surveillance in real time at the wound site. These bandages integrate smart sensors to detect the condition of wound infection based on various parameters, such as pH, temperature and oxygen level in the wound which reduces the frequency of changing the wound dressings and its associated complications. These devices can continually monitor the healing process, paving the way for tailored therapy and improved quality of patient's life. In this review, we present an overview of recent advances in biosensors for real time monitoring of pH, temperature, and oxygen in chronic wounds in order to assess infection status. We have elaborated the recent progress in quantitative monitoring of several biomarkers important for assessing wounds infection status and its detection using smart biosensors. The review shows that real-time monitoring of wound status by quantifying specific biomarkers, such as pH, temperature and tissue oxygenation to significantly aid the treatment and care of chronic infected wounds.

Microfluidic Droplet-Generation Device with Flexible Walls.

Controlling droplet sizes is one of the most important aspects of droplet generators used in biomedical research, drug discovery, high-throughput screening, and emulsion manufacturing applications. This is usually achieved by using multiple devices that are restricted in their range of generated droplet sizes. In this paper, a co-flow microfluidic droplet-generation device with flexible walls was developed such that the width of the continuous (C)-phase channel around the dispersed (D)-phase droplet-generating needle can be adjusted on demand. This actuation mechanism allowed for the adjustment of the C-phase flow velocity, hence providing modulated viscous forces to manipulate droplet sizes in a single device. Two distinct droplet-generation regimes were observed at low D-phase Weber numbers, i.e., a dripping regime at high- and medium-channel widths and a plug regime at low-channel widths. The effect of channel width on droplet size was investigated in the dripping regime under three modes of constant C-phase flow rate, velocity, and Capillary number. Reducing the channel width at a constant C-phase flow rate had the most pronounced effect on producing smaller droplets. This effect can be attributed to the combined influences of the wall effect and increased C-phase velocity, leading to a greater impact on droplet size due to the intensified viscous force. Droplet sizes in the range of 175-913 µm were generated; this range was ~2.5 times wider than the state of the art, notably using a single microfluidic device. Lastly, an empirical model based on Buckingham's Pi theorem was developed to predict the size of droplets based on channel width and height as well as the C-phase Capillary and Reynolds numbers.

Simultaneous screening of zebrafish larvae cardiac and respiratory functions: a microfluidic multi-phenotypic approach.

Multi-phenotypic screening of multiple zebrafish larvae plays an important role in enhancing the quality and speed of biological assays. Many microfluidic platforms have been presented for zebrafish phenotypic assays, but multi-organ screening of multiple larvae, from different needed orientations, in a single device that can enable rapid and large-sample testing is yet to be achieved. Here, we propose a multi-phenotypic quadruple-fish microfluidic chip for simultaneous monitoring of heart activity and fin movement of 5-7-day postfertilization zebrafish larvae trapped in the chip. In each experiment, fin movements of four larvae were quantified in the dorsal view in terms of fin beat frequency (FBF). Positioning of four optical prisms next to the traps provided the lateral views of the four larvae and enabled heart rate (HR) monitoring. The device's functionality in chemical testing was validated by assessing the impacts of ethanol on heart and fin activities. Larvae treated with 3% ethanol displayed a significant drop of 13.2 and 35.8% in HR and FBF, respectively. Subsequent tests with cadmium chloride highlighted the novel application of our device for screening the effect of heavy metals on cardiac and respiratory function at the same time. Exposure to 5 $\mu$g/l cadmium chloride revealed a significant increase of 8.2% and 39.2% in HR and FBF, respectively. The device can be employed to monitor multi-phenotypic behavioral responses of zebrafish larvae induced by chemical stimuli in various chemical screening assays, in applications such as ecotoxicology and drug discovery.

An in-vivo microfluidic assay reveals cardiac toxicity of heavy metals and the protective effect of metal responsive transcription factor (MTF-1) in Drosophila model.

Previous toxicity assessments of heavy metals on Drosophila are limited to investigating the survival, development rate, and climbing behaviour by oral administration while cardiac toxicity of these elements have not been investigated. We utilized a microfluidic device to inject known dosages of zinc (Zn) or cadmium (Cd) into the larvae's hemolymph to expose their heart directly and study their heart rate and arrhythmicity. The effect of heart-specific overexpression of metal responsive transcription factor (MTF-1) on different heartbeat parameters and survival of Drosophila larvae was investigated. The heart rate of wild-type larvae decreased by 24.8% or increased by 11.9%, 15 min after injection of 40 nL of 100 mM Zn or 10 mM Cd solution, respectively. The arrhythmicity index of wild-type larvae increased by 58.2% or 76.8%, after injection of Zn or Cd, respectively. MTF-1 heart overexpression ameliorated these effects completely. Moreover, it increased larvae's survival to pupal and adulthood stages and prolonged the longevity of flies injected with Zn and Cd. Our microfluidic-based cardiac toxicity assay illustrated that heart is an acute target of heavy metals toxicity, and MTF-1 overexpression in this tissue can ameliorate cardiac toxicity of Zn and Cd. The method can be used for cardiotoxicity assays with other pollutants in the future. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03336-7.

Low-Cost Resistive Microfluidic Salinity Sensor for High-Precision Detection of Drinking Water Salt Levels.

Rapid, inexpensive, and precise water salinity testing remains indispensable in water quality monitoring applications. Despite many sensors and commercialized devices to monitor seawater salinity, salt detection and quantification at very low levels of drinking water (below 120 ppm) have been overlooked. In this paper, we report on optimization of a low-cost microfluidic sensor to measure water salinity in the range of 1-120 ppm. The proposed design employs two copper microbridge wires suspended orthogonally in a PDMS microchannel to measure salinity based on the electrical resistance between the wires. The preliminary design of the sensor microchannel with a rectangular cross-section width (w) of 900 µm and height (h) of 500 µm could measure the water salinity in the range of 1-20 ppm in less than 1 min with detection sensitivity, limit of detection (LOD), and limit of quantification (LOQ) of 17.1 ohm/ohm·cm, 0.31 ppm, and 0.37 ppm, respectively. Data from the preliminary design was used for developing and validating a numerical model which was subsequently used for parametric studies and optimization to improve the sensor's performance. The optimized design demonstrated an order of magnitude increase in sensitivity (385 ohm/ohm·cm), a 6-fold wider detection range (1-120 ppm), and a 15-fold enhancement in miniaturization of the microfluidic channel (w = 200 µm and h = 150 µm) with LOD and LOQ of 0.39 and 0.44 ppm, respectively. In the future, the sensor can be integrated into a hand-held device to remove present impediments for low-cost and ubiquitous salinity surveillance of drinking water.

Dopaminergic signaling regulates zebrafish larvae's response to electricity.

Electrical stimulation of brain or muscle activities has gained attention for studying the molecular and cellular mechanisms involved in electric-induced responses. We recently showed zebrafish's response to electricity. Here, we hypothesized that this response is affected by the dopaminergic signaling pathways. The effects of multiple dopamine agonists and antagonists on the electric response of 6 days-postfertilization zebrafish larvae were investigated using a microfluidic device with enhanced control of experimentation and throughput. All dopamine antagonists decreased locomotor activities, while dopamine agonists did not induce similar behaviors. The D2-selective dopamine agonist quinpirole enhanced the movement. Exposure to nonselective and D1-selective dopamine agonists apomorphine and SKF-81297 caused no significant change in the electric response. Exposing larvae that were pretreated with nonselective and D2-selective dopamine antagonists butaclamol and haloperidol to apomorphine and quinpirole, respectively, restored the electric locomotion. These results reveal a correlation between electric response and dopamine signaling pathway. Furthermore, they demonstrate that electric-induced zebrafish larvae locomotion can be conditioned by modulating dopamine receptor functions. Our electrofluidic assay has profound application potential for fundamental electric-induced response research and brain disorder studies especially those related to the dopamine imbalance and as a chemical screening method when investigating biological pathways and behaviors.

Designing microfluidic devices for behavioral screening of multiple zebrafish larvae.

BACKGROUND: Microfluidic devices are being used for phenotypic screening of zebrafish larvae in fundamental and pre-clinical research. A challenge for the broad use of these microfluidic devices is their low throughput, especially in behavioral assays. Previously, we introduced the tail locomotion of a semi-mobile zebrafish larva evoked on-demand with electric signal in a microfluidic device. Here, we report the lessons learned for increasing the number of specimens from one to four larvae in this device. METHODS AND RESULTS: Multiple parameters including loading and testing time per fish and loading and orientation efficiencies were refined to optimize the performance of modified designs. Flow and electric field simulations within the final device provided insight into the flow behavior and functionality of traps when compared to previous single-larva devices. Outcomes led to a new design which decreased the testing time per larva by ≈60%. Further, loading and orientation efficiencies increased by more than 80%. Critical behavioral parameters such as response duration and tail beat frequency were similar in both single and quadruple-fish devices. CONCLUSION: The developed microfluidic device has significant advantages for greater throughput and efficiency when behavioral phenotyping is required in various applications, including chemical testing in toxicology and gene screening.

Low-cost optofluidic add-on enables rapid selective plane illumination microscopy of C. elegans with a conventional wide-field microscope.

SIGNIFICANCE: Selective plane illumination microscopy (SPIM) is an emerging fluorescent imaging technique suitable for noninvasive volumetric imaging of C. elegans. These promising microscopy systems, however, are scarce in academic and research institutions due to their high cost and technical complexities. Simple and low-cost solutions that enable conversion of commonplace wide-field microscopes to rapid SPIM platforms promote widespread adoption of SPIM by biologist for studying neuronal expressions of C. elegans. AIM: We sought to develop a simple and low-cost optofluidic add-on device that enables rapid and immobilization-free volumetric SPIM imaging of C. elegans with conventional fluorescent microscopes. APPROACH: A polydimethylsiloxane (PDMS)-based device with integrated optical and fluidic elements was developed as a low-cost and miniaturized SPIM add-on for the conventional wide-field microscope. The developed optofluidic chip contained an integrated PDMS cylindrical lens for on-chip generation of the light-sheet across a microchannel. Cross-sectional SPIM images of C. elegans were continuously acquired by the native objective of microscope as worms flowed in an L-shape microchannel and through the light sheet. RESULTS: On-chip SPIM imaging of C. elegans strains demonstrated possibility of visualizing the entire neuronal system in few seconds at single-neuron resolution, with high contrast and without worm immobilization. Volumetric visualization of neuronal system from the acquired cross-sectional two-dimensional images is also demonstrated, enabling the standard microscope to acquire three-dimensional fluorescent images of C. elegans. The full-width at half-maximum width of the point spread function was measured as 1.1 and 2.4 µm in the lateral and axial directions, respectively. CONCLUSION: The developed low-cost optofluidic device is capable of continuous SPIM imaging of C. elegans model organism with a conventional fluorescent microscope, at high speed, and with single neuron resolution.

Nanopore sensors for viral particle quantification: current progress and future prospects.

Rapid, inexpensive, and laboratory-free diagnostic of viral pathogens is highly critical in controlling viral pandemics. In recent years, nanopore-based sensors have been employed to detect, identify, and classify virus particles. By tracing ionic current containing target molecules across nano-scale pores, nanopore sensors can recognize the target molecules at the single-molecule level. In the case of viruses, they enable discrimination of individual viruses and obtaining important information on the physical and chemical properties of viral particles. Despite classical benchtop virus detection methods, such as amplification techniques (e.g., PCR) or immunological assays (e.g., ELISA), that are mainly laboratory-based, expensive and time-consuming, nanopore-based sensing methods can enable low-cost and real-time point-of-care (PoC) and point-of-need (PoN) monitoring of target viruses. This review discusses the limitations of classical virus detection methods in PoN virus monitoring and then provides a comprehensive overview of nanopore sensing technology and its emerging applications in quantifying virus particles and classifying virus sub-types. Afterward, it discusses the recent progress in the field of nanopore sensing, including integrating nanopore sensors with microfabrication technology, microfluidics and artificial intelligence, which have been demonstrated to be promising in developing the next generation of low-cost and portable biosensors for the sensitive recognition of viruses and emerging pathogens.

Conventional and microfluidic methods for airborne virus isolation and detection.

With the COVID-19 pandemic, the threat of infectious diseases to public health and safety has become much more apparent. Viral, bacterial and fungal diseases have led to the loss of millions of lives, especially in the developing world. Diseases caused by airborne viruses like SARS-CoV-2 are difficult to control, as these viruses are easily transmissible and can circulate in the air for hours. To contain outbreaks of viruses such as SARS-CoV-2 and institute targeted precautions, it is important to detect them in air and understand how they infect their targets. Point-of-care (PoC) diagnostics and point-of-need (PoN) detection methods are necessary to rapidly test patient and environmental samples, so precautions can immediately be applied. Traditional benchtop detection methods such as ELISA, PCR and culture are not suitable for PoC and PoN monitoring, because they can take hours to days and require specialized equipment. Microfluidic devices can be made at low cost to perform such assays rapidly and at the PoN. They can also be integrated with air- and liquid-based sampling technologies to capture and analyze viruses from air and body fluids. Here, conventional and microfluidic virus detection methods are reviewed and compared. The use of air sampling devices to capture and concentrate viruses is discussed first, followed by a review of analysis methods such as immunoassays, RT-PCR and isothermal amplification in conventional and microfluidic platforms. This review provides an overview of the capabilities of microfluidics in virus handling and detection, which will be useful to infectious disease researchers, biomedical engineers, and public health agencies.

High-speed label-free confocal microscopy of Caenorhabditis elegans with near infrared spectrally encoded confocal microscopy.

Caenorhabditis elegans (C. elegans) is an optically transparent nematode that shares many gene orthologs and homologs with humans. C. elegans are widely used in large populations for genetic studies relevant to human biology and disease. Success of such studies frequently relies on the ability to image C. elegans structure at high-resolution and high-speed. In this manuscript, we report on the feasibility and suitability of a high-speed variant of reflectance confocal microscopy, known as spectrally encoded confocal microscopy (SECM), for label-free imaging of C. elegans. The developed system utilizes near-infrared illumination in conjunction with refractive and diffractive optics to instantaneously image a confocal image line at a speed of up to 147 kHz with lateral and axial resolutions of 2µm and 10µm, respectively. Our imaging results from wild-type C. elegans and four mutant strains (MT2124, MT1082, CB61, and CB648) demonstrate the ability of SECM in revealing the overall geometry, key internal organs, and mutation-induced structural variations, opening the door for downstream integration of SECM in microfluidic platforms for high throughput structural imaging of C. elegans.