Single-step measurement of cell-free DNA for sepsis prognosis using a thread-based microfluidic device.

Cell-free DNA (cfDNA) content in plasma has been studied as a biomarker for sepsis. Recent publications show that the cfDNA content in sepsis patients entering intensive care unit who were likely to survive had a total cfDNA concentration of 1.16 ± 0.13 µg/mL compared to 4.65 ± 0.48 µg/mL of non-survivors. Current methods for measuring cfDNA content in plasma were designed to amplify and measure low concentrations of specific DNA, making them unsuitable for low-cost measurement of total cfDNA content in plasma. Here, we have developed a point of care (POC) device that uses a thread silicone device as a medium to store a fluorescent dye which eliminates the need for preparatory steps, external aliquoting and dispensing of reagents, preconcentration, and external mixing while reducing the detection cost. The device was paired with a portable imaging system with an excitation filter at 472 ± 10 nm and an emission filter of 520 ± 10 nm that can be operated with just 100 mA current supply. The device was demonstrated for use in the quantification of buffered cfDNA samples in a range 1-6 µg/mL with a sensitivity of 5.72 AU/µg/mL and with cfDNA spiked in plasma with a range of 1-3 µg/mL and a sensitivity of 5.43 AU/µg/mL. The results showed that the device could be used as a low-cost, rapid, and portable POC device for differentiating between survivors and non-survivors of sepsis within 20 min.

Handheld isothermal amplification and electrochemical detection of DNA in resource-limited settings.

This paper demonstrates a new method for electrochemical detection of specific sequences of DNA present in trace amounts in serum or blood. This method is designed for use at the point-of-care (particularly in resource-limited settings). By combining recombinase polymerase amplification (RPA)- an isothermal alternative to the polymerase chain reaction - with an electroactive mediator, this electrochemical methodology enables accurate detection of DNA in the field using a low-cost, portable electrochemical analyzer (specifically designed for this type of analysis). This handheld device has four attributes: (1) It uses disposable, paper-based strips that incorporate screen-printed carbon electrodes; (2) It accomplishes thermoregulation with ±0.1 °C temperature accuracy; (3) It enables electrochemical detection using a variety of pulse sequences, including square-wave and cyclic voltammetry, and coulometry; (4) It is operationally simple to use. Detection of genomic DNA from Mycobacterium smegmatis (a surrogate for M. tuberculosis-the main cause of tuberculosis), and from M. tuberculosis itself down to ∼0.040 ng/µL provides a proof-of-concept for the applicability of this method of screening for disease using molecular diagnostics. With minor modifications to the reagents, this method will also enable field monitoring of food and water quality.

Low-Cost Biosensor Technologies for Rapid Detection of COVID-19 and Future Pandemics.

Many systems have been designed for the detection of SARS-CoV-2, which is the virus that causes COVID-19. SARS-CoV-2 is readily transmitted, resulting in the rapid spread of disease in human populations. Frequent testing at the point of care (POC) is a key aspect for controlling outbreaks caused by SARS-CoV-2 and other emerging pathogens, as the early identification of infected individuals can then be followed by appropriate measures of isolation or treatment, maximizing the chances of recovery and preventing infectious spread. Diagnostic tools used for high-frequency testing should be inexpensive, provide a rapid diagnostic response without sophisticated equipment, and be amenable to manufacturing on a large scale. The application of these devices should enable large-scale data collection, help control viral transmission, and prevent disease propagation. Here we review functional nanomaterial-based optical and electrochemical biosensors for accessible POC testing for COVID-19. These biosensors incorporate nanomaterials coupled with paper-based analytical devices and other inexpensive substrates, traditional lateral flow technology (antigen and antibody immunoassays), and innovative biosensing methods. We critically discuss the advantages and disadvantages of nanobiosensor-based approaches compared to widely used technologies such as PCR, ELISA, and LAMP. Moreover, we delineate the main technological, (bio)chemical, translational, and regulatory challenges associated with developing functional and reliable biosensors, which have prevented their translation into the clinic. Finally, we highlight how nanobiosensors, given their unique advantages over existing diagnostic tests, may help in future pandemics.

Identification and characterisation of Staphylococcus aureus on low cost screen printed carbon electrodes using impedance spectroscopy.

Staphylococcus aureus infections are a cause of significant morbidity and mortality, in addition to representing a considerable economic burden. The aim of this study was to explore a low cost screen printed electrode as a sensor for the detection of S. aureus using impedance spectroscopy. S. aureus was incubated in chambers containing the electrodes and the results analysed using a novel normalisation approach. These results show that it is possible to detect the presence of S. aureus in LB media after 30 min incubation of a 1% growth culture, in addition to being able to see immediate cell concentration dependant changes in 0.9% NaCl. These observations imply that a number of electrochemical mechanisms cause a change in the impedance as a result of the presence of S. aureus, including adsorption to the electrode surface and the metabolism of the bacteria during growth. The study suggests that this detection approach would be useful in a number of clinical scenarios where S. aureus leads to difficult to treat infections.

A simple method to produce 2D and 3D microfluidic paper-based analytical devices for clinical analysis.

This paper describes the fabrication of 2D and 3D microfluidic paper-based analytical devices (µPADs) for monitoring glucose, total protein, and nitrite in blood serum and artificial urine. A new method of cutting and sealing filter paper to construct µPADs was demonstrated. Using an inexpensive home cutter printer soft cellulose-based filter paper was easily and precisely cut to produce pattern hydrophilic microchannels. 2D and 3D µPADs were designed with three detection zones each for the colorimetric detection of the analytes. A small volume of samples was added to the µPADs, which was photographed after 15 min using a digital camera. Both µPADs presented an excellent analytical performance for all analytes. The 2D device was applied in artificial urine samples and reached limits of detection (LODs) of 0.54 mM, 5.19 µM, and 2.34 µM for glucose, protein, and nitrite, respectively. The corresponding LODs of the 3D device applied for detecting the same analytes in artificial blood serum were 0.44 mM, 1.26 µM, and 4.35 µM.

Miniaturized paper-based gene sensor for rapid and sensitive identification of contagious plant virus.

Plant viruses cause significant production and economic losses in the agricultural industry worldwide. Rapid and early identification of contagious plant viruses is an essential prerequisite for the effective control of further spreading of infection. In this work, we describe a miniaturized paper-based gene sensor for the rapid and sensitive identification of a contagious plant virus. Our approach makes use of hybridization-mediated target capture based on a miniaturized lateral flow platform and gold nanoparticle colorimetric probes. The captured colorimetric probes on the test line and control line of the gene sensor produce characteristic red bands, enabling visual detection of the amplified products within minutes without the need for sophisticated instruments or the multiple incubation and washing steps performed in most other assays. Quantitative analysis is realized by recording the optical intensity of the test line. The sensor was used successfully for the identification of banana bunchy top virus (BBTV). The detection limit was 0.13 aM of gene segment, which is 10 times higher than that of electrophoresis and provides confirmation of the amplified products. We believe that this simple, rapid, and sensitive bioactive platform has great promise for warning against plant diseases in agricultural production.