Abstract
Mobile Biosensing
Integrated Quantum Dot Barcode Smartphone Optical Device for Wireless Multiplexed Diagnosis of Infected Patients
Inorganic nanoparticles are ideal precursors for engineering barcodes for rapidly detecting diseases. Despite advances in the chemical design of these barcodes, they have not advanced to clinical use because they lack sensitivity and are not cost-effective due to the need for a large readout system.
Ming et al. combine recent advances in quantum dot barcode technology with smartphones and isothermal amplification to engineer a simple and low-cost chip-based wireless multiplex diagnostic device. The authors characterize the analytical performance of this device and demonstrate that the device is capable of detecting down to 1000 viral genetic copies per milliliter, which makes it possible for clinical diagnosis of HIV or hepatitis B. More importantly, the barcoding makes it possible to detect multiple infectious pathogens simultaneously in a single test in less than 1 h.
This multiplexing capability of the device enables the diagnosis of infections that are difficult to differentiate clinically due to common symptoms such as a fever or rash. The integration of quantum dot barcoding technology with a smartphone reader provides a capacity for global surveillance of infectious diseases. (Ming, K. et al., ACS Nano.
A Smartphone Dongle for Diagnosis of Infectious Diseases at the Point-of-Care
Laksanasopin et al. demonstrate that a full laboratory-quality immunoassay can be run on a smartphone accessory. The authors report a low-cost dongle that replicates all mechanical, optical, and electronic functions of a laboratory-based enzyme-linked immunosorbent assay (ELISA) without requiring any stored energy. All necessary power is drawn from a smartphone. The dongle is placed in clinical tests in Rwanda, where health care workers use the dongle to test whole blood obtained via fingerprick from 96 patients enrolling into care at prevention of mother-to-child transmission clinics or voluntary counseling and testing centers.
The dongle performs a triplexed immunoassay not currently available in a single-test format: HIV antibody, treponemal-specific antibody for syphilis, and nontreponemal antibody for active syphilis infection. In a blinded experiment, diagnostic results obtained in 15 min from the triplex test rival the gold standard of laboratory-based HIV ELISA and rapid plasma reagin (a screening test for syphilis), with sensitivity of 92 to 100% and specificity of 79 to 100%, consistent with the needs of current clinical algorithms. Patient preference for the dongle is 97% compared to laboratory-based tests, with most pointing to the convenience of obtaining quick results with a single fingerprick.
This work suggests that coupling microfluidics with recent advances in consumer electronics can make certain laboratory-based diagnostics accessible to almost any population with access to smartphones. (Laksanasopin, T. et al., Sci Transl Med.
Rapid Imaging, Detection, and Quantification of Giardia lamblia Cysts Using Mobile Phone–Based Fluorescent Microscopy and Machine Learning
Rapid and sensitive detection of waterborne pathogens in drinkable and recreational water sources is crucial for treating and preventing the spread of water-related diseases, especially in resource-limited settings. Koydemir et al. present a field-portable and cost-effective platform for detection and quantification of Giardia lamblia cysts, one of the most common waterborne parasites, which has a thick cell wall that makes it resistant to most water disinfection techniques, including chlorination.
The platform consists of a smartphone coupled with an opto-mechanical attachment weighing ~205 g, which uses a handheld fluorescence microscope design aligned with the camera unit of the smartphone to image custom-designed disposable water sample cassettes. Each sample cassette is composed of absorbent pads and mechanical filter membranes; a membrane with 8 µm pore size is used as a porous spacing layer to prevent the backflow of particles to the upper membrane, whereas the top membrane with 5 µm pore size is used to capture the individual Giardia cysts that are fluorescently labeled.
A fluorescence image of the filter surface (field of view: ~0.8 cm2) is captured and wirelessly transmitted via the mobile phone to our servers for rapid processing using a machine learning algorithm that is trained on statistical features of Giardia cysts to automatically detect and count the cysts captured on the membrane. The results are then transmitted back to the mobile phone in less than 2 min and are displayed through a smart application running on the phone.
This mobile platform, along with a custom-developed sample preparation protocol, enables analysis of large volumes of water (e.g., 10–20 mL) for automated detection and enumeration of Giardia cysts in ~1 h, including all the steps of sample preparation and analysis. The performance of this device is evaluated using flow-cytometer-enumerated Giardia-contaminated water samples. The device demonstrates an average cyst capture efficiency of ~79% on the filter membrane along with a machine-learning-based cyst-counting sensitivity of ~84%, yielding a limit of detection of ~12 cysts per 10 mL. With its rapid detection and quantification of microorganisms, the authors believe this field-portable imaging and sensing platform running on a mobile phone could be useful for water quality monitoring in field and resource-limited settings. (Koydemir, H. C. et al., Lab Chip.
Portable Smartphone Quantitation of Prostate Specific Antigen (PSA) in a Fluoropolymer Microfluidic Device
Barbosa et al. present a new, power-free, and flexible detection system named MCFphone for portable colorimetric and fluorescence quantitative sandwich immunoassay detection of prostate specific antigen (PSA). The MCFphone is composed of a smartphone integrated with a magnifying lens, a simple light source, and a miniaturized immunoassay platform, the Microcapillary Film (MCF).
The excellent transparency and flat geometry of fluoropolymer MCF allow quantitation of PSA in the range of 0.9 to 60 ng/ml with <7% precision in 13 min using enzymatic amplification and a chromogenic substrate. The lower limit of detection is further improved from 0.4 to 0.08 ng/ml in whole-blood samples with the use of a fluorescence substrate. The MCFphone shows capability of performing rapid (13 to 22 min total assay time) colorimetric quantitative and highly sensitive fluorescence tests with good recovery rate, which represents a major step in the integration of a new generation of inexpensive and portable microfluidic devices with commercial immunoassay reagents and off-the-shelf smartphone technology. (Barbosa, A. E. et al., Bios Bioelec.
A 3D-Printed Device for a Smartphone-Based Chemiluminescence Biosensor for Lactate in Oral Fluid and Sweat
Roda et al. report a 3D-printed disposable minicartridge that can be easily prototyped to turn any kind of smartphone or tablet into a portable luminometer to detect chemiluminescence derived from enzyme-coupled reactions. As proof-of-principle, lactate oxidase is coupled with horseradish peroxidase for lactate determination in oral fluid and sweat. Lactate can be quantified in less than 5 min with detection limits of 0.5 mmol L−1 and 0.1 mmol L−1 in oral fluid and sweat, respectively. This smartphone-based device shows adequate analytical performance to offer a cost-effective alternative for noninvasive lactate measurement. The authors believe it can be used to evaluate lactate variation in relation to the anaerobic threshold in endurance sport and for monitoring lactic acidosis in critical-care patients. (Roda, A. et al., Analyst.
Paper and Printed Biosensors
Paper-Based Inkjet-Printed Microfluidic Analytical Devices
Rapid, precise, and reproducible deposition of a broad variety of functional materials, including analytical assay reagents and biomolecules, has made inkjet printing an effective tool for the fabrication of microanalytical devices. A ubiquitous office device as simple as a standard desktop printer with its multiple ink cartridges can be used for this purpose.
Yamada et al. publish a review discussing the combination of inkjet printing technology with paper as a printing substrate for the fabrication of microfluidic paper-based analytical devices (µPADs), which have developed into a fast-growing new field in analytical chemistry. After introducing the fundamentals of µPADs and inkjet printing, the review touches on topics such as the microfluidic patterning of paper, tailored arrangement of materials, and functionalities achievable exclusively by the inkjet deposition of analytical assay components, before concluding with an outlook on future perspectives. (Yamada, K. et al., Angew Chem. 54,
A Low-Cost and Simple Paper-Based Microfluidic Device for Simultaneous Multiplex Determination of Different Types of Chemical Contaminants in Food
It is difficult to carry out multiple detection of different types of chemicals because of different chemical microenvironment requirements. Zhang et al. report a low-cost and simple paper-based microfluidic device integrated with a fluorescence-labeled single-stranded DNA (ssDNA) functionalized graphene oxide sensor to address the needs of multiplex detection of different types of chemical contaminants in food.
In this work, Cy5-labeled corresponding functional ssDNA for different analytes associated with graphene oxide (ssDNA-GO) are used as core detection sensors to sensitively report the presence of different types of chemicals as well as enlarge chemical compatibility, which makes it possible to simultaneously detect multiple chemicals in the same chemical microenvironment. This paper microfluidic device can be fabricated easily, and the paper substrate also facilitates the integration of ssDNA-GO sensors via physical absorption. This device is successfully applied in multiplex detection of heavy metal mercury (II) ion (Hg2+) and silver (I) ion (Ag+) and aminoglycoside antibiotics residues in food, and also provides great potential for applications in environmental monitoring and clinical diagnosis. (Zhang, Y. et al., Bios Bioelec.
A Stacking Flow Immunoassay for the Detection of Dengue-Specific Immunoglobulins in Salivary Fluid
Paper-based immunoassays, usually in the form of lateral flow tests, are currently the standard platform for home diagnostics. However, conventional lateral tests are often complicated by severe nonspecific adsorption of detector particles when applied to test samples containing salivary fluid. It is believed that a high concentration of proteinaceous substances in salivary fluid causes particle aggregation and adhesion.
In a recent study, Zhang et al. develop a stacking flow platform for single-step detection of a target antibody in salivary fluid. Stacking flow circumvents the need for separate sample pretreatments, such as filtration or centrifugation, which are often required prior to testing saliva samples when using paper-based immunoassays. This is achieved by guiding the samples and reagents to the test strip through different paths. By doing so, salivary substances that interfere with the particle-based sensing system are removed before they come into contact with the detection reagents, which greatly reduces the background. In addition, the stacking flow configuration enables uniform flow with a unique flow regulator, which leads to even test lines with good quantification capability, enabling the detection of ~20 ng mL−1 α-fetoprotein in the serum. This stacking flow device has been successfully applied to detect dengue-specific immunoglobulins that are present in salivary fluid. (Zhang, Y. et al., Lab Chip.
Reagent Pencils: A New Technique for Solvent-Free Deposition of Reagents onto Paper-Based Microfluidic Devices
Mitchell et al. report custom-made pencils containing reagents dispersed in a solid matrix to enable rapid and solvent-free deposition of reagents onto membrane-based fluidic devices. The technique is as simple as drawing with the reagent pencils on a device. When aqueous samples are added to the device, the reagents dissolve from the pencil matrix and become available to react with analytes in the sample.
Colorimetric glucose assays conducted on devices prepared using reagent pencils have comparable accuracy and precision to assays conducted on conventional devices prepared with reagents deposited from solution. Most importantly, sensitive reagents, such as enzymes, are stable in the pencils under ambient conditions, and no significant decrease in the activity of the enzyme horseradish peroxidase stored in a pencil was observed after 63 days. The authors believe reagent pencils offer a new option for preparing and customizing diagnostic tests at the point-of-care without the need for specialized equipment. (Mitchell, H. T. et al., Lab Chip.
New Biosensing Mechanisms
Large-Area CMOS Bio-Pixel Array for Compact High-Sensitive Multiplex Biosensing
Sandeau et al. demonstrate a novel complementary metal oxide semiconductor (CMOS) bio-pixel array that integrates assay substrate and assay readout for multiplex and multireplicate detection of a triplicate of cytokines with single-digit pg ml−1 sensitivities. The device features large-area bio-pixels that enable individual assays to be dedicated to and addressed by single pixels. The device contains 128 available pixels that enable simultaneous measurement of a large number of targets.
Chemiluminescent assays are carried out directly on the pixel surfaces, and the emitted chemiluminescent photons are subsequently registered by pixels, making possible a highly compact sensor and reader format. The high sensitivity of the bio-pixel array is enabled by the high refractive index of silicon-based pixels. This in turn generates a strong supercritical angle luminescence response, significantly increasing the efficiency of the photon collection over conventional far-field modalities. (Sandeau, L. et al., Lab Chip.
Performance Characterization of an Abiotic and Fluorescent-Based Continuous Glucose-Monitoring System in Patients with Type 1 Diabetes
Mortellaro and DeHennis report a continuous glucose-monitoring (CGM) system consisting of a wireless, subcutaneously implantable glucose sensor and a body-worn transmitter, and conduct a clinical evaluation over a 28-day implant period in 12 type 1 diabetic patients. The implantable sensor is constructed of a fluorescent, boronic-acid-based, glucose-indicating polymer coated onto a miniaturized, polymer-encased optical detection system. The external transmitter wirelessly communicates with and powers the sensor, and it contains Bluetooth capability for interfacing with a smartphone application. The accuracy of 19 implanted sensors is evaluated over 28 days during six in-clinic sessions by comparing the CGM glucose values to venous blood glucose measurements taken every 15 min. Mean absolute relative difference (MARD) for all sensors is 11.6±0.7%, and Clarke error grid analysis shows that 99% of paired data points are in the combined A and B zones. (Mortellaro, M. and DeHennis, A., Biosens Bioelectron.
Metabolic Profiling of Bacteria by Unnatural C-Terminated D-Amino Acids
Bacterial peptidoglycan is a mesh-like network composed of sugars and oligopeptides. Transpeptidases cross-link peptidoglycan oligopeptides to provide vital cell wall rigidity and structural support. It was recently discovered that the same transpeptidases catalyze the metabolic incorporation of exogenous D-amino acids onto bacterial cell surfaces with vast promiscuity for the side-chain identity.
Pidgeon et al. report that this enzymatic promiscuity is not exclusive to side chains, but that C-terminus variations can also be accommodated among a diverse range of bacteria. Atomic force microscopy analysis reveals that the incorporation of C-terminus amidated D-amino acids onto bacterial surfaces substantially reduces the cell wall stiffness. The authors exploit the promiscuity of bacterial transpeptidases to develop a novel assay for profiling different bacterial species. (Pidgeon, S. E. et al., Angew Chem.
Direct, Rapid, and Label-Free Detection of Enzyme–Substrate Interactions in Physiological Buffers Using CMOS-Compatible Nanoribbon Sensors
Mu et al. explore the use of Al2O3-passivated Si nanowire devices (nanoribbons) in the analysis of enzyme–substrate interactions via the monitoring of pH change. The authors demonstrate the effectiveness of the device through the detection of urea in phosphate buffered saline (PBS), and penicillinase in PBS and urine, at limits of detection of <200 µM and 0.02 units/mL, respectively, and demonstrate the device’s capability to extract accurate enzyme kinetics. The Michaelis–Menten constant (Km) from the acetylcholine–acetylcholinesterase reaction is also demonstrated. (Mu, L. et al., Nano Lett.
Mediatorless Glucose Biosensor and Direct Electron Transfer Type Glucose/Air Biofuel Cell Enabled with Carbon Nanodots
Zhao et al. report the application of carbon nanodots (CNDs), a new type of carbonaceous nanomaterials, in electrochemical sensing. In this study, CNDs are used as immobilization supports and electron carriers to promote direct electron transfer (DET) reactions of glucose oxidase (GOx) and bilirubin oxidase (BOD). In a CND electrode modified with GOx, a high rate constant (ks) of 6.28±0.05 s–1 for fast DET and an apparent Michaelis–Menten constant (KMapp) as low as 0.85±0.03 mM for affinity to glucose are found.
By taking advantage of its excellent direct bioelectrocatalytic performances with glucose oxidation, a DET-based biosensor for glucose detection ranging from 0 to 0.64 mM with a high sensitivity of 6.1 µA mM–1 and a limit of detection (LOD) of 1.07±0.03 µM (S/N = 3) is achieved. In addition, the promoted DET of BOD immobilized on CNDs is shown to effectively catalyze the reduction of oxygen to water at the onset potential of +0.51 V (vs Ag/AgCl). A mediator-free DET-type glucose/air enzymatic biofuel cell (BFC), in which CND electrodes modified with GOx and BOD are used for oxidizing glucose at the bioanode and reducing oxygen at the biocathode, respectively, is fabricated. The BFC displays an open-circuit voltage (OCV) as high as 0.93 V and a maximum power density of 40.8 µW cm–2 at 0.41 V. (Zhao, M. et al., Anal Chem.
Portable and Visual Electrochemical Sensor Based on the Bipolar Light-Emitting Diode Electrode
Zhang et al. report a novel sensing strategy based on a closed bipolar system consisting of a light-emitting diode (LED) and a split bipolar electrode (BPE) for generation of luminescent signal in the presence of the target. With this design, a BPE array is constructed for the quick and high-throughput determination of various electroactive substances with the naked eye. Due to the ultrahigh current efficiency of the closed bipolar system, the sample concentration can be reported by the luminous intensity of the inserted LED without the expensive luminescent agent and instruments. In addition, the stability of the signal is improved because of the electroluminescent property of the LED. The authors demonstrate applications of this bipolar LED electrode (BP-LED-E) with rapid quantification of four model targets: H2O2, ascorbic acid, glucose, and blood sugar. (Zhang, X. et al., Anal Chem.
Electrochemical Assay to Detect Influenza Viruses and Measure Drug Susceptibility
Zhang et al. report an electrochemical assay for rapid diagnosis of influenza viruses. Glucose-bearing substrate is exposed to influenza viruses’ enzyme neuraminidase (NA). The enzyme causes the release of glucose, which is detected amperometrically.
Two methods are used to detect released glucose. First, the authors use a standard glucose blood meter to detect two viral NAs and three influenza strains. With this method, the authors also demonstrate drug susceptibility of two antivirals, Zanamivir and Oseltamivir, which can be measured. In addition, the authors also use disposable test strips to detect 19 H1N1 and H3N2 influenza strains in 1 h, and they find the limit and range of detection of this assay to be 102 and 102–108 plaque-forming units, respectively. (Zhang, X. et al., Angew Chem.