Photonic-Plasmonic Coupling Enhanced Fluorescence Enabling Digital-Resolution Ultrasensitive Protein Detection.

Assays utilizing fluorophores are common throughout life science research and diagnostics, although detection limits are generally limited by weak emission intensity, thus requiring many labeled target molecules to combine their output to achieve higher signal-to-noise. We describe how the synergistic coupling of plasmonic and photonic modes can significantly boost the emission from fluorophores. By optimally matching the resonant modes of a plasmonic fluor (PF) nanoparticle and a photonic crystal (PC) with the absorption and emission spectrum of the fluorescent dye, a 52-fold improvement in signal intensity is observed, enabling individual PFs to be observed and digitally counted, where one PF tag represents one detected target molecule. The amplification can be attributed to the strong near-field enhancement due to the cavity-induced activation of the PF, PC band structure-mediated improvement in collection efficiency, and increased rate of spontaneous emission. The applicability of the method by dose-response characterization of a sandwich immunoassay for human interleukin-6, a biomarker used to assist diagnosis of cancer, inflammation, sepsis, and autoimmune disease is demonstrated. A limit of detection of 10 fg mL-1 and 100 fg mL-1 in buffer and human plasma respectively, is achieved, representing a capability nearly three orders of magnitude lower than standard immunoassays.

A photonic resonator interferometric scattering microscope for label-free detection of nanometer-scale objects with digital precision in point-of-use environments.

Label-free detection and digital counting of nanometer-scaled objects such as nanoparticles, viruses, extracellular vesicles, and protein molecules enable a wide range of applications in cancer diagnostics, pathogen detection, and life science research. Here, we report the design, implementation, and characterization of a compact Photonic Resonator Interferometric Scattering Microscope (PRISM) designed for point-of-use environments and applications. The contrast of interferometric scattering microscopy is amplified through a photonic crystal surface, upon which scattered light from an object combines with illumination from a monochromatic source. The use of a photonic crystal substrate for interferemetric scattering microscopy results in reduced requirements for high-intensity lasers or oil-immersion objectives, thus opening a pathway toward instruments that are more suitable for environments outside the optics laboratory. The instrument incorporates two innovative elements that facilitate operation on a desktop in ordinary laboratory environments by users that do not have optics expertise. First, because scattering microscopes are extremely sensitive to vibration, we incorporated an inexpensive but effective solution of suspending the instrument's main components from a rigid metal framework using elastic bands, resulting in an average of 28.7 dBV reduction in vibration amplitude compared to an office desk. Second, an automated focusing module based on the principle of total internal reflection maintains the stability of image contrast over time and spatial position. In this work, we characterize the system's performance by measuring the contrast from gold nanoparticles with diameters in the 10-40 nm range and by observing various biological analytes, including HIV virus, SARS-CoV-2 virus, exosome, and ferritin protein.

Rapid detection of intact SARS-CoV-2 using designer DNA Nets and a pocket-size smartphone-linked fluorimeter.

Rapid, sensitive, and inexpensive point-of-care diagnosis is vital to controlling highly infectious diseases, including COVID-19. Here, we report the design and characterization of a compact fluorimeter called a "Virus Pod" (V-Pod) that enables sensitive self-testing of SARS-CoV-2 viral load in saliva. The rechargeable battery-operated device reads the fluorescence generated by Designer DNA Nanostructures (DDN) when they specifically interact with intact SARS-CoV-2 virions. DDNs are net-shaped self-assembling nucleic acid constructs that provide an array of highly specific aptamer-fluorescent quencher duplexes located at precise positions that match the pattern of spike proteins. The room-temperature assay is performed by mixing the test sample with DNA Net sensor in a conventional PCR tube and placing the tube into the V-Pod. Fluorescent signals are generated when multivalent aptamer-spike binding releases fluorescent quenchers, resulting in rapid (5-min) generation of dose-dependent output. The V-Pod instrument performs laser excitation, fluorescence intensity quantitation, and secure transmission of data to an App via Bluetooth™. We show that the V-Pod and DNA Net assay achieves clinically relevant detection limits of 3.92 × 103 viral-genome-copies/mL for pseudo-typed wild-type SARS-CoV-2 and 1.84 × 104, 9.69 × 104, 6.99 × 104 viral-genome-copies/mL for pathogenic Delta, Omicron, and D614G variants, representing sensitivity similar to laboratory-based PCR. The pocket-sized instrument (∼$294), inexpensive reagent-cost/test ($1.26), single-step, rapid sample-to-answer, and quantitative output represent a capability that is compatible with the needs of frequent self-testing in a consumer-friendly format that can link with medical service systems such as healthcare providers, contact tracing, and infectious disease reporting.

Review of HIV Self Testing Technologies and Promising Approaches for the Next Generation.

The ability to self-test for HIV is vital to preventing transmission, particularly when used in concert with HIV biomedical prevention modalities, such as pre-exposure prophylaxis (PrEP). In this paper, we review recent developments in HIV self-testing and self-sampling methods, and the potential future impact of novel materials and methods that emerged through efforts to develop more effective point-of-care (POC) SARS-CoV-2 diagnostics. We address the gaps in existing HIV self-testing technologies, where improvements in test sensitivity, sample-to-answer time, simplicity, and cost are needed to enhance diagnostic accuracy and widespread accessibility. We discuss potential paths toward the next generation of HIV self-testing through sample collection materials, biosensing assay techniques, and miniaturized instrumentation. We discuss the implications for other applications, such as self-monitoring of HIV viral load and other infectious diseases.

A Photonic Resonator Interferometric Scattering Microscope for Label-free Detection of Nanometer-Scale Objects with Digital Precision in Point-of-Use Environments.

Label-free detection and digital counting of nanometer-scaled objects such as nanoparticles, viruses, extracellular vesicles, and protein molecules enable a wide range of applications in cancer diagnostics, pathogen detection, and life science research. The contrast of interferometric scattering microscopy is amplified through a photonic crystal surface, upon which scattered light from an object combines with illumination from a monochromatic plane wave source. The use of a photonic crystal substrate for interference scattering microscopy results in reduced requirements for high-intensity lasers or oil-immersion objectives, thus opening a pathway toward instruments that are more suitable for environments outside the optics laboratory. Here, we report the design, implementation, and characterization of a compact Photonic Resonator Interferometric Scattering Microscope (PRISM) designed for point-of-use environments and applications. The instrument incorporates two innovative elements that facilitate operation on a desktop in ordinary laboratory environments by users that do not have optics expertise. First, because scattering microscopes are extremely sensitive to vibration, we incorporated an inexpensive but effective solution of suspending the instrument's main components from a rigid metal framework using elastic bands, resulting in an average of 28.7 dBV reduction in vibration amplitude compared to an office desk. Second, an automated focusing module based on the principle of total internal reflection maintains the stability of image contrast over time and spatial position, facilitating automated data collection. In this work, we characterize the system's performance by measuring the contrast from gold nanoparticles with diameters in the 10-40 nm range and by observing various biological analytes, including HIV virus, SARS-CoV-2 virus, exosomes, and ferritin protein.

Smartphone clip-on instrument and microfluidic processor for rapid sample-to-answer detection of Zika virus in whole blood using spatial RT-LAMP.

Rapid, simple, inexpensive, accurate, and sensitive point-of-care (POC) detection of viral pathogens in bodily fluids is a vital component of controlling the spread of infectious diseases. The predominant laboratory-based methods for sample processing and nucleic acid detection face limitations that prevent them from gaining wide adoption for POC applications in low-resource settings and self-testing scenarios. Here, we report the design and characterization of an integrated system for rapid sample-to-answer detection of a viral pathogen in a droplet of whole blood comprised of a 2-stage microfluidic cartridge for sample processing and nucleic acid amplification, and a clip-on detection instrument that interfaces with the image sensor of a smartphone. The cartridge is designed to release viral RNA from Zika virus in whole blood using chemical lysis, followed by mixing with the assay buffer for performing reverse-transcriptase loop-mediated isothermal amplification (RT-LAMP) reactions in six parallel microfluidic compartments. The battery-powered handheld detection instrument uniformly heats the compartments from below, and an array of LEDs illuminates from above, while the generation of fluorescent reporters in the compartments is kinetically monitored by collecting a series of smartphone images. We characterize the assay time and detection limits for detecting Zika RNA and gamma ray-deactivated Zika virus spiked into buffer and whole blood and compare the performance of the same assay when conducted in conventional PCR tubes. Our approach for kinetic monitoring of the fluorescence-generating process in the microfluidic compartments enables spatial analysis of early fluorescent "bloom" events for positive samples, in an approach called "Spatial LAMP" (S-LAMP). We show that S-LAMP image analysis reduces the time required to designate an assay as a positive test, compared to conventional analysis of the average fluorescent intensity of the entire compartment. S-LAMP enables the RT-LAMP process to be as short as 22 minutes, resulting in a total sample-to-answer time in the range of 17-32 minutes to distinguish positive from negative samples, while demonstrating a viral RNA detection as low as 2.70 × 102 copies per µl, and a gamma-irradiated virus of 103 virus particles in a single 12.5 µl droplet blood sample.

A paper-based cantilever array sensor: Monitoring volatile organic compounds with naked eye.

Volatile organic compound (VOC) detection is critical for controlling industrial and commercial emissions, environmental monitoring, and public health. Simple, portable, rapid and low-cost VOC sensing platforms offer the benefits of on-site and real-time monitoring anytime and anywhere. The best and most practically useful approaches to monitoring would include equipment-free and power-free detection by the naked eye. In this work, we created a novel, paper-based cantilever sensor array that allows simple and rapid naked-eye VOC detection without the need for power, electronics or readout interface/equipment. This simple VOC detection method was achieved using (i) low-cost paper materials as a substrate and (ii) swellable thin polymers adhered to the paper. Upon exposure to VOCs, the polymer swelling adhered to the paper-based cantilever, inducing mechanical deflection that generated a distinctive composite pattern of the deflection angles for a specific VOC. The angle is directly measured by the naked eye on a 3-D protractor printed on a paper facing the cantilevers. The generated angle patterns are subjected to statistical algorithms (linear discriminant analysis (LDA)) to classify each VOC sample and selectively detect a VOC. We classified four VOC samples with 100% accuracy using LDA.

A micro-sized bio-solar cell for self-sustaining power generation.

Self-sustainable energy sources are essential for a wide array of wireless applications deployed in remote field locations. Due to their self-assembling and self-repairing properties, "biological solar (bio-solar) cells" are recently gaining attention for those applications. The bio-solar cell can continuously generate electricity from microbial photosynthetic and respiratory activities under day-night cycles. Despite the vast potential and promise of bio-solar cells, they, however, have not yet successfully been translated into commercial applications, as they possess persistent performance limitations and scale-up bottlenecks. Here, we report an entirely self-sustainable and scalable microliter-sized bio-solar cell with significant power enhancement by maximizing solar energy capture, bacterial attachment, and air bubble volume in well-controlled microchambers. The bio-solar cell has a ~300 µL single chamber defined by laser-machined poly(methyl methacrylate) (PMMA) substrates and it uses an air cathode to allow freely available oxygen to act as an electron acceptor. We generated a maximum power density of 0.9 mW m(-2) through photosynthetic reactions of cyanobacteria, Synechocystis sp. PCC 6803, which is the highest power density among all micro-sized bio-solar cells.