Serodiagnosis of leishmaniasis in asymptomatic and symptomatic dogs by use of the recombinant dynamin-1-like protein from Leishmania infantum: A preliminary study.

Visceral leishmaniasis (VL) is a fatal manifestation of an infection caused by intracellular protozoa of the Leishmania genus. In New World countries, VL is classified as a zoonotic disease with domestic dogs acting as its main reservoir. Asymptomatic dogs are as competent to transmit Leishmania to the vectors as symptomatic dogs, however current diagnostic tests are limited and present low sensitivity for this important group. The development of accurate tests is fundamental to the early diagnosis, treatment, and control of canine leishmaniasis. In this study, we investigated the use of a recombinant protein (dynamin-1-like protein, Dyn-1) from L. infantum, as a potential target antigen for leishmaniasis serodiagnosis in both symptomatic and asymptomatic dogs. The antigenic performance of the protein was evaluated by means of ELISA assays using sera from symptomatic (n = 25), asymptomatic (n = 34) and non-infected dogs (n = 36) using ELISA. In addition, sera from dogs experimentally infected with Trypanosoma cruzi (n = 49) and naturally infected with Babesia sp. (n = 8) were tested to evaluate possible cross-reactivity. A crude soluble antigen (CSA) of Leishmania was used as an antigen control and K39 and K26 were used as reference antigens because they are already widely used in commercial tests. rDyn-1-based assay showed the highest sensitivity (97%) compared to the antigens K39 (88%), K26 (86%) and crude extract (95%). The highest specificity among the tests was also obtained with the protein rDyn-1 (94%), compared with the other antigens K39 (81%), K26 (87%), and crude extract (77%). This study showed that the rDyn-1 ELISA assay was able to identify 100% of asymptomatic dogs, establishing its potential as a target for the diagnosis of canine leishmaniasis.

Thermal Detection of Glucose in Urine Using a Molecularly Imprinted Polymer as a Recognition Element.

Glucose bio-sensing technologies have received increasing attention in the last few decades, primarily due to the fundamental role that glucose metabolism plays in diseases (e.g., diabetes). Molecularly imprinted polymers (MIPs) could offer an alternative means of analysis to a field that is traditionally dominated by enzyme-based devices, posing superior chemical stability, cost-effectiveness, and ease of fabrication. Their integration into sensing devices as recognition elements has been extensively studied with different readout methods such as quartz-crystal microbalance or impedance spectroscopy. In this work, a dummy imprinting approach is introduced, describing the synthesis and optimization of a MIP toward the sensing of glucose. Integration of this polymer into a thermally conductive receptor layer was achieved by micro-contact deposition. In essence, the MIP particles are pressed into a polyvinyl chloride adhesive layer using a polydimethylsiloxane stamp. The prepared layer is then evaluated with the so-called heat-transfer method, allowing the determination of the specificity and the sensitivity of the receptor layer. Furthermore, the selectivity was assessed by analyzing the thermal response after infusion with increasing concentrations of different saccharide analogues in phosphate-buffered saline (PBS). The obtained results show a linear range of the sensor of 0.0194-0.3300 mM for the detection of glucose in PBS. Finally, a potential application of the sensor was demonstrated by exposing the receptor layer to increasing concentrations of glucose in human urine samples, demonstrating a linear range of 0.0444-0.3300 mM. The results obtained in this paper highlight the applicability of the sensor both in terms of non-invasive glucose monitoring and for the analysis of food samples.

Topographical Vacuum Sealing of 3D-Printed Multiplanar Microfluidic Structures.

We demonstrate a novel way of creating three-dimensional microfluidic channels capable of following complex topographies. To this end, substrates with open channels and different geometries were 3D-printed, and the open channels were consecutively closed with a thermoplastic using a low-resolution vacuum-forming approach. This process allows the sealing of channels that are located on the surface of complex multiplanar topographies, as the thermoplastic aligns with the surface-shape (the macrostructure) of the substrate, while the microchannels remain mostly free of thermoplastic as their small channel size resists thermoplastic inflow. This new process was analyzed for its capability to consistently close different substrate geometries, which showed reliable sealing of angles >90°. Furthermore, the thermoplastic intrusion into channels of different widths was quantified, showing a linear effect of channel width and percentage of thermoplastic intrusion; ranging from 43.76% for large channels with 2 mm width to only 5.33% for channels with 500 µm channel width. The challenging sealing of substrate 'valleys', which are created when two large protrusions are adjacent to each other, was investigated and the correlation between protrusion distance and height is shown. Lastly, we present three application examples: a serpentine mixer with channels spun around a cuboid, increasing the usable surface area; a cuvette-inspired flow cell for a 2-MXP biosensor based on molecular imprinted polymers, fitting inside a standard UV/Vis-Spectrophotometer; and an adapter system that can be manufactured by one-sided injection molding and is self-sealed before usage. These examples demonstrate how this novel technology can be used to easily adapt microfluidic circuits for application in biosensor platforms.

Modular Science Kit as a support platform for STEM learning in primary and secondary school.

The need to develop interest in STEM (science, technology, engineering, and mathematics) skills in young pupils has driven many educational systems to include STEM as a subject in primary schools. In this work, a science kit aimed at children from 8 to 14 years old is presented as a support platform for an innovative and stimulating approach to STEM learning. The peculiar design of the kit, based on modular components, is aimed to help develop a multitude of skills in the young students, dividing the learning process into two phases. During phase 1 the pupils build the experimental setup and visualize the scientific phenomena, while in phase 2, they are introduced and challenged to understand the principles on which these phenomena are based, guided by a handbook. This approach aims at making the experience more inclusive, stimulating the interest and passion of the pupils for scientific subjects.

A Molecularly Imprinted Polymer-based Dye Displacement Assay for the Rapid Visual Detection of Amphetamine in Urine.

The rapid sensing of drug compounds has traditionally relied on antibodies, enzymes and electrochemical reactions. These technologies can frequently produce false positives/negatives and require specific conditions to operate. Akin to antibodies, molecularly imprinted polymers (MIPs) are a more robust synthetic alternative with the ability to bind a target molecule with an affinity comparable to that of its natural counterparts. With this in mind, the research presented in this article introduces a facile MIP-based dye displacement assay for the detection of (±) amphetamine in urine. The selective nature of MIPs coupled with a displaceable dye enables the resulting low-cost assay to rapidly produce a clear visual confirmation of a target's presence, offering huge commercial potential. The following manuscript characterizes the proposed assay, drawing attention to various facets of the sensor design and optimization. To this end, synthesis of a MIP tailored towards amphetamine is described, scrutinizing the composition and selectivity (ibuprofen, naproxen, 2-methoxphenidine, quetiapine) of the reported synthetic receptor. Dye selection for the development of the displacement assay follows, proceeded by optimization of the displacement process by investigating the time taken and the amount of MIP powder required for optimum displacement. An optimized dose-response curve is then presented, introducing (±) amphetamine hydrochloride (0.01-1 mg mL-1) to the engineered sensor and determining the limit of detection (LoD). The research culminates in the assay being used for the analysis of spiked urine samples (amphetamine, ibuprofen, naproxen, 2-methoxphenidine, quetiapine, bupropion, pheniramine, bromopheniramine) and evaluating its potential as a low-cost, rapid and selective method of analysis.

Point of Care Diagnostics in Resource-Limited Settings: A Review of the Present and Future of PoC in Its Most Needed Environment.

Point of care (PoC) diagnostics are at the focus of government initiatives, NGOs and fundamental research alike. In high-income countries, the hope is to streamline the diagnostic procedure, minimize costs and make healthcare processes more efficient and faster, which, in some cases, can be more a matter of convenience than necessity. However, in resource-limited settings such as low-income countries, PoC-diagnostics might be the only viable route, when the next laboratory is hours away. Therefore, it is especially important to focus research into novel diagnostics for these countries in order to alleviate suffering due to infectious disease. In this review, the current research describing the use of PoC diagnostics in resource-limited settings and the potential bottlenecks along the value chain that prevent their widespread application is summarized. To this end, we will look at literature that investigates different parts of the value chain, such as fundamental research and market economics, as well as actual use at healthcare providers. We aim to create an integrated picture of potential PoC barriers, from the first start of research at universities to patient treatment in the field. Results from the literature will be discussed with the aim to bring all important steps and aspects together in order to illustrate how effectively PoC is being used in low-income countries. In addition, we discuss what is needed to improve the situation further, in order to use this technology to its fullest advantage and avoid "leaks in the pipeline", when a promising device fails to take the next step of the valorization pathway and is abandoned.

Rapid Colorimetric Screening of Elevated Phosphate in Urine: A Charge-Transfer Interaction.

A charge-transfer (CT) interaction between 1,3,5-trinitro-2,4-dimethylbenzene (TNX) and anionic phosphate is evaluated, yielding a high band electronic transfer interaction that can be observed as a distinct color change when phosphate is present in solution. The induced interaction was studied using 1H NMR, UV-visible, and Fourier transform infrared spectroscopies. The stoichiometric determination of the interaction was divined by means of continuous variation, applying the Schaeppi-Treadwell method to calculate the binding constant (k). Furthermore, the effect of the polarity of solvents toward the generation of the CT interaction was examined, with multiple solvents considered. Complex deconstruction studies were undertaken, examining the effects of water on complex destruction and understanding the volumes needed to hinder the CT interaction potency. Specificity and selectivity of the CT interaction were also studied against other biologically relevant species (CH3CH2OH, Na+, K+, Ca2+, Cl-, HCO3 -, F-, CH3COO-, and SO4 2-), assessing the capabilities of the assay to differentiate anionic species and counter cations that could act as interferences. The role of TNX concentration in CT formation was also analyzed, aiming to optimize the phosphate-sensing assay and improve its limit of detection. The sensing platform was subsequently used to study phosphate concentrations in urine samples to further understand its potential application in biomedical research. To validate the developed technique, urine samples were analyzed for their phosphate content with both the developed sensor and a validated vanadate-molybdate reagent. The results indicate that the sensing method is capable of accurately reporting elevated phosphate levels in urine samples in a rapid and sensitive manner, illustrating that the colorimetric test could be used as a prescreening test for conditions such as hyperphosphatemia or chronic kidney disease.

A Novel Biomimetic Tool for Assessing Vitamin K Status Based on Molecularly Imprinted Polymers.

Vitamin K was originally discovered as a cofactor required to activate clotting factors and has recently been shown to play a key role in the regulation of soft tissue calcification. This property of vitamin K has led to an increased interest in novel methods for accurate vitamin K detection. Molecularly Imprinted Polymers (MIPs) could offer a solution, as they have been used as synthetic receptors in a large variety of biomimetic sensors for the detection of similar molecules over the past few decades, because of their robust nature and remarkable selectivity. In this article, the authors introduce a novel imprinting approach to create a MIP that is able to selectively rebind vitamin K1. As the native structure of the vitamin does not allow for imprinting, an alternative imprinting strategy was developed, using the synthetic compound menadione (vitamin K3) as a template. Target rebinding was analyzed by means of UV-visible (UV-VIS) spectroscopy and two custom-made thermal readout techniques. This analysis reveals that the MIP-based sensor reacts to an increasing concentration of both menadione and vitamin K1. The Limit of Detection (LoD) for both compounds was established at 700 nM for the Heat Transfer Method (HTM), while the optimized readout approach, Thermal Wave Transport Analysis (TWTA), displayed an increased sensitivity with a LoD of 200 nM. The sensor seems to react to a lesser extent to Vitamin E, the analogue under study. To further demonstrate its potential application in biochemical research, the sensor was used to measure the absorption of vitamin K in blood serum after taking vitamin K supplements. By employing a gradual enrichment strategy, the sensor was able to detect the difference between baseline and peak absorption samples and was able to quantify the vitamin K concentration in good agreement with a validation experiment using High-Performance Liquid Chromatography (HPLC). In this way, the authors provide a first proof of principle for a low-cost, straightforward, and label-free vitamin K sensor.