Disposable Peptidoglycan-Specific Biosensor for Noninvasive Real-Time Detection of Broad-Spectrum Gram-Positive Bacteria in Exhaled Breath Condensates.

Rapidly identifying and quantifying Gram-positive bacteria are crucial to diagnosing and treating bacterial lower respiratory tract infections (LRTIs). This work presents a field-deployable biosensor for detecting Gram-positive bacteria from exhaled breath condensates (EBCs) based on peptidoglycan recognition using an aptamer. Dielectrophoretic force is employed to enrich the bacteria in 10 s without additional equipment or steps. Concurrently, the measurement of the sensor's interfacial capacitance is coupled to quantify the bacteria during the enrichment process. By incorporation of a semiconductor condenser, the whole detection process, including EBC collection, takes about 3 min. This biosensor has a detection limit of 10 CFU/mL, a linear range of up to 105 CFU/mL and a selectivity of 1479:1. It is cost-effective and disposable due to its low cost. The sensor provides a nonstaining, culture-free and PCR-independent solution for noninvasive and real-time diagnosis of Gram-positive bacterial LRTIs.

Capacitive Aptasensor Coupled with Microfluidic Enrichment for Real-Time Detection of Trace SARS-CoV-2 Nucleocapsid Protein.

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has lasted for almost 2 years. Stemming its spread has posed severe challenges for clinical virus detection. A long turnaround time, complicated operation, and low accuracy have become bottlenecks in developing detection techniques. Adopting a direct antigen detection strategy, we developed a fast-responding and quantitative capacitive aptasensor for ultratrace nucleocapsid protein detection based on a low-cost microelectrode array (MEA) chip. Employing the solid-liquid interface capacitance with a sensitivity of picofarad level, the tiny change on the MEA surface can be definitively detected. As a result, the limit of detection reaches an ultralow level of femtogram per milliliter in different matrices. Integrated with efficient microfluidic enrichment, the response time of this sensor from the sample to the result is shortened to 15 s, completely meeting the real-time detection demand. Moreover, the wide linear range of the sensor is from 10-5 to 10-2 ng/mL, and a high selectivity of 6369:1 is achieved. After application and evaluation in different environmental and body fluid matrices, this sensor and the detection method have proved to be a label-free, real-time, easy-to-operate, and specific strategy for SARS-CoV-2 screening and diagnosis.

Numerical investigation of microchannel geometry for effective on-chip biofluid delivery by AC electrothermal effect.

Although there exist tremendous needs for on-chip biofluid delivery, research in this field has yielded limited numbers of devices for real-world applications. One challenge is the difficulty for micropumps to meet the requirements of being low cost to fabricate, easy to integrate and effective for intended applications at the same time. This research focuses on AC electrothermal (ACET) micropumps based on planar interdigitated electrodes, due to their practicality in fabrication and operation, and compatibility with biochemical fluids. Our prior work has optimized the design of electrode dimensions for a fixed microchannel design. This work finds that microchannel dimensions can also affect ACET micropumps significantly, with respect to flow rate and electric impedance loading. This work first considers the constraints arising from impedance loading by ACET micropumps on power supplies, then the investigation describes several key parameters (threshold height, saturation thickness), to arrive at an appropriate microchannel geometry for the effective delivery of biofluids. The optimized microchannel is expected to incorporate well into a multifunctional lab-chip system to transport biofluids efficiently.

An alternating current electrokinetics biosensor for rapid on-site serological screening of Taenia solium cysticercosis infection.

Cysticercosis, caused by Taenia solium infection, is a leading cause of acquired epilepsy in many developing countries. Several types of immunoassays have been developed for the detection of Taenia solium infection in both infected humans and livestock animals. However, these methods require central laboratory facilities and are both time- and labor-consuming with longer than desired turnaround time. In this work, we demonstrated that AC electrokinetics (ACEK) capacitive sensing can be used to realize point-of-care immunosensor in general, with the on-site screening of Taenia solium infection as an example here. The sensor employs interdigitated microelectrodes (IDME) functionalized with a recombinant Taenia solium antigen, rT24H, to detect anti-rT24H antibodies in clinical serum samples. ACEK capacitive sensing method interrogates the IDME sensors with a special AC signal, which serves the dual purposes of enriching target antibodies by ACEK effects and directly measuring the capacitance change induced by specific binding. First, to characterize the ACEK biosensor as an immunosensor in general, IgG in phosphate-buffered saline buffer was tested against IDME sensors functionalized with anti-IgG. The limit of detection of the sensor was 24.1 fg/mL, and the linear dynamic range was 0.1-100 pg/mL. To test the clinical usage of this sensor, ACEK capacitive sensors with rT24H probe were used to test clinical serum samples from patients with or without Taenia solium infection. The diagnostic sensitivity of the ACEK capacitive sensor for Taenia solium infection was found to be 88.24%. ACEK capacitive immunosensors have shown good potential for point-of-care diagnostics.

A Sensitive and Specific Genomic RNA Sensor for Point-of-Care Screening of Zika Virus from Serum.

This work presents a sensitive and specific single-step RNA sensor for Zika virus (ZIKV) in serum. Using AC electrokinetics (ACEK)-enhanced capacitive sensing technology, ZIKV genomic RNA (gRNA) can be directly detected from serum. The sensors are interdigitated electrodes modified with oligonucleotide probes complementary to the conserved regions of ZIKV gRNA. The ACEK capacitive sensing applies an optimized AC excitation signal over the sensor, which induces ACEK microfluidic enrichment of analytes and also simultaneously performs real-time monitoring of hybridization of ZIKV gRNA on the sensor surface. Hence, the sensing procedures are simple with rapid turn-around time and good specificity and sensitivity. A series of experiments are conducted to optimize the sensor performance. The performance of the sensor is investigated for three different probes, two functionalization buffers, and different hybridization buffers. With the optimized sensing protocol, this method can detect spiked ZIKV gRNA from human serum within 30 s and reach a limit of detection of 78.8 copies/µL in analytical samples and as low as 287.5 copies/µL in neat serum. The sensors can successfully differentiate between the RNAs of the ZIKV and dengue virus, two viruses with similar transmission paths and symptoms. The sensor is simple to use and requires no labeling or sophisticated process typically involved in a polymerase chain reaction, hybridization chain reaction, or nucleic acid sequence-based amplification.

AC electrokinetic drug delivery in dentistry using an interdigitated electrode assembly powered by inductive coupling.

AC electrokinetics (ACEK) has been shown to deliver certain drugs into human teeth more effectively than diffusion. However, using electrical wires to power intraoral ACEK devices poses risks to patients. The study demonstrates a novel interdigitated electrode arrays (IDE) assembly powered by inductive coupling to induce ACEK effects at appropriate frequencies to motivate drugs wirelessly. A signal generator produces the modulating signal, which multiplies with the carrier signal to produce the amplitude modulated (AM) signal. The AM signal goes through the inductive link to appear on the secondary coil, then rectified and filtered to dispose of its carrier signal, and the positive half of the modulating signal appears on the load. After characterizing the device, the device is validated under light microscopy by motivating carboxylate-modified microspheres, tetracycline, acetaminophen, benzocaine, lidocaine and carbamide peroxide particles with induced ACEK effects. The assembly is finally tested in a common dental bleaching application. After applying 35 % carbamide peroxide to human teeth topically or with the IDE at 1200 Hz, 5 Vpp for 20 min, spectrophotometric analysis showed that compared to diffusion, the IDE enhanced whitening in specular optic and specular optic excluded modes by 215 % and 194 % respectively. Carbamide peroxide absorbance by the ACEK group was two times greater than diffusion as measured by colorimetric oxidation-reduction and UV-Vis spectroscopy at 550 nm. The device motivates drugs of variable molecular weight and structure wirelessly. Wireless transport of drugs to intraoral targets under ACEK effects may potentially improve the efficacy and safety of drug delivery in dentistry.

Photothermal-Based Multiplex Nested Digital PCR System for Rapid Detection of Foodborne Pathogens.

The rapid and sensitive detection of foodborne pathogens is crucial for ensuring food safety. Among virus testing methods, polymerase chain reaction (PCR) has served as the gold-standard technique in most food safety regulation organizations. However, to enhance the speed and efficiency of PCR, novel approaches are continually being explored. In this work, leveraging the photothermal effects and high thermal conductivity of gold nanoparticles, we have significantly improved the heating and cooling rates of thermal cycles, enabling ultra-fast PCR detection. Specifically, we present a pre-degassing multiplex digital PCR chip integrated with gold nanoparticles. We further developed a portable system with a light source for photothermal heating cycling, along with an optoelectronic sensor to analyze PCR amplification products after rapid thermal cycling. As proof of concept, the proposed chip and portable device was applied for the on-site detection of several types of foodborne pathogens, including Escherichia coli, Listeria monocytogenes, Staphylococcus aureus, and Salmonella. The whole system could distinguish those pathogens within 20 min, showing good potential for the rapid detection of multiple types of foodborne pathogens.

Virtual Multiplexing Chamber-Based Digital PCR for Camel Milk Authentication Applications.

In this work, we proposed a chamber-based digital PCR (cdPCR) microfluidic device that is compatible with fluorescence imaging systems for milk adulteration detection. The device enables the digitalization of PCR reagents, which are loaded into microchambers, and subsequent thermocycling for DNA amplification. Then, fluorescence images of the microchambers are captured and analyzed to obtain the total number of positive chambers, which is used to calculate the copy numbers of the target DNA, enabling accurate quantitative detections to determine intentional milk adulteration from accidental contaminations. The validation of this device is performed by camel milk authentication. We performed 25,600-chamber virtual multiplexing cdPCR tests using 40 × 40 chamber devices for the detection of DNA templates extracted from pure or mixed milk with different dilutions. Then, the cdPCR chip was used to authenticate blind milk samples, demonstrating its efficacy in real biotechnical applications.

Label-Free Sensing of Cell Viability Using a Low-Cost Impedance Cytometry Device.

Cell viability is an essential physiological status for drug screening. While cell staining is a conventional cell viability analysis method, dye staining is usually cytotoxic. Alternatively, impedance cytometry provides a straightforward and label-free sensing approach for the assessment of cell viability. A key element of impedance cytometry is its sensing electrodes. Most state-of-the-art electrodes are made of expensive metals, microfabricated by lithography, with a typical size of ten microns. In this work, we proposed a low-cost microfluidic impedance cytometry device with 100-micron wide indium tin oxide (ITO) electrodes to achieve a comparable performance to the 10-micron wide Au electrodes. The effectiveness was experimentally verified as 7 µm beads can be distinguished from 10 µm beads. To the best of our knowledge, this is the lowest geometry ratio of the target to the sensing unit in the impedance cytometry technology. Furthermore, a cell viability test was performed on MCF-7 cells. The proposed double differential impedance cytometry device has successfully differentiated the living and dead MCF-7 cells with a throughput of ~1000 cells/s. The label-free and low-cost, high-throughput impedance cytometry could benefit drug screening, fundamental biological research and other biomedical applications.

Nucleic Acid Detection with Ion Concentration Polarization Microfluidic Chip for Reduced Cycle Numbers of Polymerase Chain Reaction.

Nucleic acid detection is widely used in disease diagnosis, food safety, environmental monitoring and many other research fields. The continuous development of rapid and sensitive new methods to detective nucleic acid is very important for practical application. In this study, we developed a rapid nucleic-acid detection method using polymerase chain reaction (PCR) combined with electrokinetic preconcentration based on ion concentration polarization (ICP). Using a Nafion film, the proposed ICP microfluidic chip is utilized to enrich the nucleic acid molecules amplified by PCR thermal cycles. To demonstrate the capability of the microfluidic device and the hybrid nucleic-acid detection method, we present an animal-derived component detection experiment for meat product identification applications. With the reduced cycle numbers of 24 cycles, the detection can be completed in about 35 min. The experimental results show that this work can provide a microfluidic device and straightforward method for rapid detection of nucleic acids with reduced cycle numbers.

Microfluidics-Integrated Sensors toward Rapid Detection of Single Nucleotide Variations.

Rapid detection of single nucleotide variations (SNVs) is of critical importance to early diagnosis of several diseases and the prediction of diverse responses to a specific treatment. Based on the information published in the literature, discrimination of SNVs is a developing area of study with great research enthusiasm and is also an area that can benefit from microfluidics-integrated designs. This review provides a brief overview of different microfluidics-based strategies for rapid detection of SNVs and mismatched bases. Sensors based on various microfluidic formats, such as paper-based microfluidic biosensors, droplet-based microfluidic systems, and magnetic bead-based microfluidic biosensors, have been discussed with respect to their specific pros and cons for SNV detection. These systems have shown promise for distributed on-site diagnostics in personalized medicine.