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The analysis and detection of snake venom toxins are a matter of great importance in clinical diagnosis for fast treatment and the discovery of new pharmaceutical products. Current detection methods have high associated costs and require the use of sophisticated bioreceptors, which in some cases are difficult to obtain. Herein, we report the synthesis of template-based molecularly imprinted micromotors for dynamic detection of α-bungarotoxin as a model toxin present in the venom of many-banded krait (Bungarus multicinctus). The specific recognition sites are built-in in the micromotors by incubation of the membrane template with the target toxin, followed by a controlled electrodeposition of a poly(3,4-ethylenedioxythiophene)/poly(sodium 4-styrenesulfonate) polymeric layer, a magnetic Ni layer to promote magnetic guidance and facilitate washing steps, and a Pt layer for autonomous propulsion in the presence of hydrogen peroxide. The enhanced fluid mixing and autonomous propulsion increase the likelihood of interactions with the target analyte as compared with static counterparts, retaining the tetramethylrhodamine-labeled α-bungarotoxin on the micromotor surface with extremely fast dynamic sensor response (after just 20 s navigation) in only 3 µL of water, urine, or serum samples. The sensitivity achieved meets the clinically relevant concentration postsnakebite (from 0.1 to 100 µg/mL), illustrating the feasibility of the approach for practical applications. The selectivity of the protocol is very high, as illustrated by the absence of fluorescence in the micromotor surface in the presence of α-cobratoxin as a representative toxin with a size and structure similar to those of α-bungarotoxin. Recoveries higher than 95% are obtained in the analysis of urine- and serum-fortified samples. The new strategy holds considerable promise for fast, inexpensive, and even onsite detection of several toxins using multiple molecularly imprinted micromotors with tailored recognition abilities.
Assuntos
Bungarotoxinas , Bungarotoxinas/química , Bungarotoxinas/urina , Animais , Polímeros/química , Venenos de Serpentes/química , Bungarus , Compostos Bicíclicos Heterocíclicos com Pontes/química , Impressão Molecular , Ácidos SulfônicosRESUMO
Micromotor (MM) technology offers a valuable and smart on-the-move biosensing microscale approach in clinical settings where sample availability is scarce in the case of Alzheimer's disease (AD). Soluble amyloid-ß protein oligomers (AßO) (mainly AßO42) that circulate in biological fluids have been recognized as a molecular biomarker and therapeutic target of AD due to their high toxicity, and they are correlated much more strongly with AD compared to the insoluble Aß monomers. A graphene oxide (GO)-gold nanoparticles (AuNPs)/nickel (Ni)/platinum nanoparticles (PtNPs) micromotors (MMGO-AuNPs)-based electrochemical label-free aptassay is proposed for sensitive, accurate, and rapid determination of AßO42 in complex clinical samples such as brain tissue, cerebrospinal fluid (CSF), and plasma from AD patients. An approach that implies the in situ formation of AuNPs on the GO external layer of tubular MM in only one step during MM electrosynthesis was performed (MMGO-AuNPs). The AßO42 specific thiolated-aptamer (AptAßO42) was immobilized in the MMGO-AuNPs via Au-S interaction, allowing for the selective recognition of the AßO42 (MMGO-AuNPs-AptAßO42-AßO42). AuNPs were smartly used not only to covalently bind a specific thiolated-aptamer for the design of a label-free electrochemical aptassay but also to improve the final MM propulsion performance due to their catalytic activity (approximately 2.0× speed). This on-the-move bioplatform provided a fast (5 min), selective, precise (RSD < 8%), and accurate quantification of AßO42 (recoveries 94-102%) with excellent sensitivity (LOD = 0.10 pg mL-1) and wide linear range (0.5-500 pg mL-1) in ultralow volumes of the clinical sample of AD patients (5 µL), without any dilution. Remarkably, our MM-based bioplatform demonstrated the competitiveness for the determination of AßO42 in the target samples against the dot blot analysis, which requires more than 14 h to provide qualitative results only. It is also important to highlight its applicability to the potential analysis of liquid biopsies as plasma and CSF samples, improving the reliability of the diagnosis given the heterogeneity and temporal complexity of neurodegenerative diseases. The excellent results obtained demonstrate the analytical potency of our approach as a future tool for clinical/POCT (Point-of-care testing) routine scenarios.
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Doença de Alzheimer , Técnicas Biossensoriais , Grafite , Nanopartículas Metálicas , Humanos , Doença de Alzheimer/diagnóstico , Doença de Alzheimer/metabolismo , Ouro/química , Peptídeos beta-Amiloides/análise , Nanopartículas Metálicas/química , Reprodutibilidade dos Testes , Limite de Detecção , Platina , Proteínas Amiloidogênicas , Técnicas Biossensoriais/métodos , Técnicas Eletroquímicas/métodosRESUMO
3D printing technology is a tremendously powerful technology to fabricate electrochemical sensing devices. However, current conductive filaments are not aimed at electrochemical applications and therefore require intense activation protocols to unleash a suitable electrochemical performance. Current activation methods based on (electro)chemical activation (using strong alkaline solutions and organic solvents and/or electrochemical treatments) or combined approaches are time-consuming and require hazardous chemicals and dedicated operator intervention. Here, pioneering spark-discharge-activated 3D-printed electrodes were developed and characterized, and it was demonstrated that their electrochemical performance was greatly improved by the effective removal of the thermoplastic support polylactic acid (PLA) as well as the formation of sponge-like and low-dimensional carbon nanostructures. This reagent-free approach consists of a direct, fast, and automatized spark discharge between the 3D-electrode and the respective graphite pencil electrode tip using a high-voltage power supply. Activated electrodes were challenged toward the simultaneous voltammetric determination of dopamine (DP) and serotonin (5-HT) in cell culture media. Spark discharge has been demonstrated as a promising approach for conductive filament activation as it is a fast, green (0.94 GREEnness Metric Approach), and automatized procedure that can be integrated into the 3D printing pipeline.
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Given the long-life expectancy of the newborn, research aimed at improving sepsis diagnosis and management in this population has been recognized as cost-effective, which at early stages continues to be a tremendous challenge. Despite there is not an ideal-specific biomarker, the simultaneous detection of biomarkers with different behavior during an infection such as procalcitonin (PCT) as high specificity biomarker with one of the earliest biomarkers in sepsis as interleukin-6 (IL-6) increases diagnostic performance. This is not only due to their high positive predictive value but also, since it can also help the clinician to rule out infection and thus avoid the use of antibiotics, due to their high negative predictive value. To this end, we explore a cutting-edge micromotor (MM)-based OFF-ON dual aptassay for simultaneous determination of both biomarkers in 15 min using just 2 µL of sample from low-birth-weight neonates with gestational age less than 32 weeks and birthweight below 1000 g with clinical suspicion of late-onset sepsis. The approach reached the high sensitivities demanded in the clinical scenario (LODPCT = 0.003 ng/mL, LODIL6 = 0.15 pg/mL) with excellent correlation performance (r > 0.9990, p < 0.05) of the MM-based approach with the Hospital method for both biomarkers during the analysis of diagnosed samples and reliability (Er < 6% for PCT, and Er < 4% for IL-6). The proposed approach also encompasses distinctive technical attributes in a clinical scenario since its minimal sample volume requirements and expeditious results compatible with few easy-to-obtain drops of heel stick blood samples from newborns admitted to the neonatal intensive care unit. This would enable the monitoring of both sepsis biomarkers within the initial hours after the manifestation of symptoms in high-risk neonates as a valuable tool in facilitating prompt and well-informed decisions about the initiation of antibiotic therapy.These results revealed the asset behind micromotor technology for multiplexing analysis in diagnosing neonatal sepsis, opening new avenues in low sample volume-based diagnostics.
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Sepse Neonatal , Sepse , Recém-Nascido , Humanos , Lactente , Sepse Neonatal/diagnóstico , Sepse Neonatal/tratamento farmacológico , Calcitonina , Proteína C-Reativa/análise , Interleucina-6 , Reprodutibilidade dos Testes , Análise Custo-Benefício , Sepse/diagnóstico , Biomarcadores , Pró-Calcitonina , Antibacterianos/uso terapêuticoRESUMO
A one-shot CO2 laser-based strategy to generate conductive reduced graphene oxide (rGO) decorated with nanoceria (nCe) is proposed. The 2D/0D rGO-nCe films, integrated as catalytic sensing layers in paper-based sensors, were employed for on-site monitoring of indoor fogging treatments against Listeria monocytogenes (Lm), a ubiquitous pathogenic bacterium. The rGO-nCe laser-assisted synthesis was optimized to preserve the rGO film morphological and electron-transfer features and simultaneously integrate catalytic nCe. The films were characterized by microscopical (SEM), spectroscopical (EDX, Raman, and FTIR), and electrochemical techniques. The most performing film was integrated into a nitrocellulose substrate, and the complete sensor was assembled via a combination of xurography and stencil printing. The rGO-nCe sensor's catalytic activity was proved toward the detection of H2O2, obtaining sensitive determination (LOD = 0.3 µM) and an extended linear range (0.5-1500 µM). Eventually, the rGO-nCe sensor was challenged for the real-time continuous monitoring of hydrogen peroxide aerosol during no-touch fogging treatment conducted following the EU's recommendation for biocidal product use. Treatment effectiveness was proved toward three Lm strains characterized by different origins, i.e., type strain ATCC 7644, clinical strain 338, and food strain 641/6II. The sensor allows for discrimination and quantification treatments at different environmental biocidal amounts and fogging times, and correlates with the microbiological inhibition, promoting the proposed sensor as a useful tool to modulate and monitor no-touch treatments.
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Desinfecção , Grafite , Peróxido de Hidrogênio , Lasers , Listeria monocytogenes , Papel , Grafite/química , Peróxido de Hidrogênio/química , Listeria monocytogenes/efeitos dos fármacos , Listeria monocytogenes/isolamento & purificação , Desinfecção/métodos , Cério/química , Limite de Detecção , Técnicas Eletroquímicas/métodos , Técnicas Eletroquímicas/instrumentação , CatáliseRESUMO
Three-dimensional (3D) printing technology has emerged as a powerful technology for the fabrication of low-cost microfluidics. Nevertheless, the fabrication of microfluidic devices integrating high-performance electrochemical sensors in practical applications is still an open challenge. Although automatic fabrication of the microfluidic device and the electrodes can be successfully carried out using a one-step multimaterial fused filament fabrication (FFF) approach, the as-printed electrochemical performance of these electrodes is not good enough for chemical (bio)sensing and their surface modification is challenging because after closing the channel there is no physical access to the electrode. Thus, here a pause-print-pause (PPP) microfabrication approach was implemented. The fabrication was paused before printing the microfluidics, and the filament-based electrodes were directly modified on the printing bed via stencil printing, drop casting, and electrodeposition. To exemplify this versatile workflow, the design of a microfluidic glucose sensor was proposed. To this end, first, the working and counter electrodes were stencil printed with graphite ink while the reference electrode was stencil printed with Ag|AgCl ink. Then, Prussian blue was formed on the working electrode either by drop casting or by electrodeposition, and glucose oxidase was drop cast on top. At this point, the microfabrication process was resumed, and the microfluidics were printed on top of the modified electrodes to complete the construction of hybrid electrochemical fluidic fused filament fabricated devices (h-eF4Ds). This print-pause-print approach is not limited to ink-based electrodes or glucose oxidase, and we envisage these results will pave the way for the effective integration of electrodes in microfluidic devices in a simple and clean-room-free approach, allowing the development of highly customized eF4Ds for a plethora of analytes with high significance.
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For the first time, a paper-based analytical device (PAD) was developed for the assessment of transferrin saturation (TSAT), which is defined as the ratio between iron bound to transferrin (Tf) and the total iron-binding capacity (TIBC) of Tf. Both parameters were simultaneously measured on the same PAD using ferrozine as a chromophore and a smartphone as the color reader. To this end, Tf was first isolated from serum using anti-Tf immunomagnetic beads to ensure that only the Tf-bound iron was measured, improving the selectivity and accuracy of TSAT assessment. To demonstrate the practical utility of the device, it was validated by analyzing a certified reference material, showing excellent accuracy (Er < 4%) and good precision (RSD ≤ 6%). Finally, 18 diagnosed serum samples from ischemic stroke patients were analyzed by this approach, and the results were compared with those obtained by urea-PAGE, showing not only an excellent correlation (r = 0.93, p < 0.05) but that the PAD approach has become statistically identical to the free-interference urea-PAGE. In comparison with the slow, tedious, and non-miniaturized-PAGE, this PAD approach exhibited attractive characteristics such as low cost, disposability, and connectivity, showing great potential for future point-of-care testing, especially in developing countries and/or remote areas, where access to medical or clinical facilities is limited.
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AVC Isquêmico , Humanos , AVC Isquêmico/diagnóstico , Ferro/sangue , Transferrinas/sangue , Cor , Aplicativos MóveisRESUMO
Miniaturized magnetic-based pipette tip microextraction is presented as a sample preparation approach for microsamples. It involves quick dispersion of a diminutive amount of a magnetic sorbent material in a low-volume sample (10 µL) to entrap the target analytes. Next, the dispersion is aspirated using a (semi)automatic pipette through a pipette tip with a small cubic neodymium magnet inside, which retrieves the magnetic sorbent containing the analytes. After discarding the rest of the sample, the sorbent is properly rinsed by aspirating/dispensing deionized water, and then, the analytes are eluted by aspirating/dispensing an appropriate solvent. This approach was employed for the determination of free cortisol in serum and urine from very low birth weight preterm newborns, a vulnerable patient group who present low availability for sampling biological fluids. A magnetic immunosorbent made of a cortisol antibody was employed for the selective extraction, followed by liquid chromatography-tandem mass spectrometry. Good analytical features were obtained, such as limits of detection and quantification of 0.08 and 0.27 ng mL-1, respectively, linearity up to 50 ng mL-1 (R2 > 0.999), RSD values under 15% and relative recoveries between 91 and 111%. The cross-reactivity with other glucocorticoids (i.e., cortisone and prednisolone) was evaluated to show the selectivity of the extraction. Finally, the method applicability was demonstrated towards the determination of free cortisol in the serum and urine samples from low birth weight preterm newborns.
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Hidrocortisona , Extração em Fase Sólida , Recém-Nascido , Humanos , Extração em Fase Sólida/métodos , Cromatografia Líquida , Recém-Nascido de muito Baixo Peso , Fenômenos Magnéticos , Limite de DetecçãoRESUMO
Prussian Blue (PB)/chitosan enzyme mimetic tubular micromotors are used here for on-the-fly (bio)-sensing assays. The micromotors are easily prepared by direct deposition of chitosan into the pores of a membrane template and in situ PB synthesis during hydrogel deposition. Under judicious pH control, PB micromotors display enzyme mimetic capabilities with three key functions on board: the autonomous oxygen bubble propulsion (with PB acting as a catalase mimic for hydrogen peroxide decomposition), 3,3',5,5'-tetramethylbenzidine (TMB) oxidation (with PB acting as a peroxidase mimic for analyte detection), and as a magnetic material (to simplify the (bio)-sensing steps). In connection with chitosan capabilities, these unique enzyme mimetic micromotors are further functionalized with acetylthiocholinesterase enzyme (ATChE) to be explored in fast inhibition assays (20 min) for the colorimetric determination of the nerve agent neostigmine, with excellent analytical performance in terms of quantification limit (0.30 µM) and concentration linear range (up to 500 µM), without compromising efficient micromotor propulsion. The new concept illustrated holds considerable potential for a myriad of (bio)-sensing applications, including forensics, where this conceptual approach remains to be explored. Micromotor-based tests to be used in crime scenes are also envisioned due to the reliable neostigmine determination in unpretreated samples.
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Quitosana , Colorimetria , Ferrocianetos , Peróxido de Hidrogênio/química , NeostigminaRESUMO
2D layered molybdenum disulfide (MoS2 ) nanomaterials are a promising platform for biomedical applications, particularly due to its high biocompatibility characteristics, mechanical and electrical properties, and flexible functionalization. Additionally, the bandgap of MoS2 can be engineered to absorb light over a wide range of wavelengths, which can then be transformed into local heat for applications in photothermal tissue ablation and regeneration. However, limitations such as poor stability of aqueous dispersions and low accumulation in affected tissues impair the full realization of MoS2 for biomedical applications. To overcome such challenges, herein, multifunctional MoS2 -based magnetic helical microrobots (MoSBOTs) using cyanobacterium Spirulina platensis are proposed as biotemplate for therapeutic and biorecognition applications. The cytocompatible microrobots combine remote magnetic navigation with MoS2 photothermal activity under near-infrared irradiation. The resulting photoabsorbent features of the MoSBOTs are exploited for targeted photothermal ablation of cancer cells and on-the-fly biorecognition in minimally invasive oncotherapy applications. The proposed multi-therapeutic MoSBOTs hold considerable potential for a myriad of cancer treatment and diagnostic-related applications, circumventing current challenges of ablative procedures.
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Molibdênio , Nanoestruturas , Dissulfetos , Raios Infravermelhos , Fototerapia/métodosRESUMO
Micro/nanomotors are nanoscale devices that have been explored in various fields, such as drug delivery, environmental remediation, or biosensing and diagnosis. The use of micro/nanomotors has grown considerably over the past few years, partially because of the advantages that they offer in the development of new conceptual avenues in biosensing. This is due to their propulsion and intermixing in solution compared with their respective static forms, which enables motion-based detection methods and/or decreases bioassay time. This review focuses on the impacts of micro/nanomotors on biosensing research in the last 2 years. An overview of designs for bioreceptor attachment to micro/nanomotors is given. Recent developments have focused on chemically propelled micromotors using external fuels, commonly hydrogen peroxide. However, the associated fuel toxicity and inconvenience of use in relevant biological samples such as blood have prompted researchers to explore new micro/nanomotor biosensing approaches based on biocompatible propulsion sources such as magnetic or ultrasound fields. The main advances in biocompatible propulsion sources for micro/nanomotors as novel biosensing platforms are discussed and grouped by their propulsion-driven forces. The relevant analytical applications are discussed and representatively illustrated. Moreover, envisioning future biosensing applications, the principal advantages of micro/nanomotor synthesis using biocompatible and biodegradable materials are given. The review concludes with a realistic drawing on the present and future perspectives.
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Recuperação e Remediação Ambiental , Nanoestruturas , Sistemas de Liberação de Medicamentos , Peróxido de Hidrogênio , Nanotecnologia/métodosRESUMO
Surface-enhanced Raman scattering (SERS) is a very promising tool for the direct detection of biomarkers for the diagnosis of i.e., cancer and pathogens. Yet, current SERS strategies are hampered by non-specific interactions with co-existing substances in the biological matrices and the difficulties of obtaining molecular fingerprint information from the complex vibrational spectrum. Raman signal enhancement is necessary, along with convenient surface modification and machine-based learning to address the former issues. This review aims to describe recent advances and prospects in SERS-based approaches for cancer and pathogens diagnosis. First, direct SERS strategies for key biomarker sensing, including the use of substrates such as plasmonic, semiconductor structures, and 3D order nanostructures for signal enhancement will be discussed. Secondly, we will illustrate recent advances for indirect diagnosis using active nanomaterials, Raman reporters, and specific capture elements as SERS tags. Thirdly, critical challenges for translating the potential of the SERS sensing techniques into clinical applications via machine learning and portable instrumentation will be described. The unique nature and integrated sensing capabilities of SERS provide great promise for early cancer diagnosis or fast pathogens detection, reducing sanitary costs but most importantly allowing disease prevention and decreasing mortality rates.
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Nanoestruturas , Análise Espectral Raman , Análise Espectral Raman/métodos , Nanoestruturas/químicaRESUMO
Janus micromotors encapsulating transition metal dichalcogenides (TMDs) and modified with a rhodamine (RhO)-labeled affinity peptide (RhO-NFMESLPRLGMH) are used here for Salmonella enterica endotoxin detection. The OFF-ON strategy relies on the specific binding of the peptide with the TMDs to induce fluorescence quenching (OFF state); which is next recovered due to selectively binding to the endotoxin (ON state). The increase in the fluorescence of the micromotors can be quantified as a function of the concentration of endotoxin in the sample. The developed strategy was applied to the determination of Salmonella enterica serovar Typhimurium endotoxin with high sensitivity (limits of detection (LODs) of 2.0 µg/mL using MoS2, and 1.2 µg/mL using WS2), with quantitative recoveries (ranging from 93.7 ± 4.6 % to 94.3 ± 6.6%) in bacteria cultures in just 5 min. No fluorescence recovery is observed in the presence of endotoxins with a similar structure, illustrating the high selectivity of the protocol, even against endotoxins of Salmonella enterica serovar Enteritidis with great similarity in its structure, demonstrating the high bacterial specificity of the developed method. These results revealed the analytical potential of the reported strategy in multiplexed assays using different receptors or in the design of portable detection devices.
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Salmonella enterica , Salmonella enteritidis , Endotoxinas , Limite de Detecção , PeptídeosRESUMO
The ubiquity and importance of ROS and RNS in cellular signaling, disease development, and death give rise to an outstanding interest in their detection and quantification. Among the analytical techniques available, electrochemical sensors stand out for the detection of ROS/RNS due to their high sensitivity and inherent miniaturization which allows the in situ and real-time detection together with a tunable selectivity due to the different electrochemical behavior of ROS/RNS. Nanomaterial-based enzyme-free electrochemical sensors possess improved sensitivity, selectivity, stability, and unique catalytic activities. In addition, their integration in nanoelectrodes, lab-on-chips, microfluidic systems, and stretchable electrodes allow the determination of ROS/RNS in individual cells, cell organelles, or cell populations, under different experimental conditions hardly accessible using classical detection methods.
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Técnicas Eletroquímicas , Espécies Reativas de Nitrogênio/análise , Espécies Reativas de Oxigênio/análise , Animais , Células Cultivadas , Humanos , Nanoestruturas/química , Espécies Reativas de Nitrogênio/metabolismo , Espécies Reativas de Oxigênio/metabolismoRESUMO
Two cellulose nanocrystals/single-walled carbon nanotube (CNC/SW) hybrids, using two cellulose polymorphs, were evaluated as electrochemical transducers: CNC type I (CNC-I/SW) and CNC type II (CNC-II/SW). They were synthesized and fully characterized, and their analytical performance as electrochemical sensors was carefully studied. In comparison with SWCNT-based and screen-printed carbon electrodes, CNC/SW sensors showed superior electroanalytical performance in terms of sensitivity and selectivity, not only in the detection of small metabolites (uric acid, dopamine, and tyrosine) but also in the detection of complex glycoproteins (alpha-1-acid glycoprotein (AGP)). More importantly, CNC-II/SW exhibited 20 times higher sensitivity than CNC-I/SW for AGP determination, yielding a LOD of 7 mg L-1.These results demonstrate the critical role played by nanocellulose polymorphism in the electrochemical performance of CNC/SW hybrid materials, opening new directions in the electrochemical sensing of these complex molecules. In general, these high-active-surface hybrids smartly exploited the preserved non-oxidized SW conductivity with the high aqueous dispersibility of the CNC, avoiding the use of organic solvents or the incorporation of toxic surfactants during their processing, making the CNC/SW hybrids promising nanomaterials for electrochemical detection following greener approaches.
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Celulose/química , Técnicas Eletroquímicas/métodos , Nanotubos de Carbono/química , Estrutura Molecular , Nanocompostos/químicaRESUMO
Herein, we describe a Janus micromotor smartphone platform for the motion-based detection of glutathione. The system compromises a universal three-dimensional (3D)-printed platform to hold a commercial smartphone, which is equipped with an external magnification optical lens (20-400×) directly attached to the camera, an adjustable sample holder to accommodate a glass slide, and a light-emitting diode (LED) source. The presence of glutathione in peroxide-rich sample media results in the decrease in the speed of 20 µm graphene-wrapped/PtNPs Janus micromotors due to poisoning of the catalytic layer by a thiol bond formation. The speed can be correlated with the concentration of glutathione, achieving a limit of detection of 0.90 µM, with percent recoveries and excellent selectivity under the presence of interfering amino acids and proteins. Naked-eye visualization of the speed decrease allows for the design of a test strip for fast glutathione detection (30 s), avoiding previous amplification strategies or sample preparation steps. The concept can be extended to other micromotor approaches relying on fluorescence or colorimetric detection for future multiplexed schemes.
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Glutationa , SmartphoneRESUMO
Janus particles are a unique type of materials combining two different functionalities in a single unit. This allows the combination of different analytical properties leading to new analytical capabilities, i.e., enhanced fluid mixing to increase sensitivity with targeting capturing abilities and unique advantages in terms of multi-functionality and versatility of modification, use, and operation both in static and dynamic modes. The aim of this conceptual review is to cover recent (over the last 5 years) advances in the use of Janus microparticles and micromotors in (bio)-sensing. First, the role of different materials and synthetic routes in the performance of Janus particles are described. In a second main section, electrochemical and optical biosensing based on Janus particles and motors are covered, including in vivo and in vitro methodologies as the next biosensing generation. Current challenges and future perspectives are provided in the conclusions section.
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Técnicas Biossensoriais/instrumentação , Nanopartículas Multifuncionais , Nanopartículas/químicaRESUMO
Graphene oxide/PtNPs/Fe2 O3 "dual-propelled" catalytic and fuel-free rotary actuated magnetic Janus micromotors modified with the lanbiotic Nisin are used for highly selective capture/inactivation of gram-positive bacteria units and biofilms. Specific interaction of Nisin with the Lipid II unit of Staphylococcus Aureus bacteria in connection with the enhanced micromotor movement and generated fluid flow result in a 2-fold increase of the capture/killing ability (both in bubble and magnetic propulsion modes) as compared with free peptide and static counterparts. The high stability of Nisin along with the high towing force of the micromotors allow for efficient operation in untreated raw media (tap water, juice and serum) and even in blood and in flowing blood in magnetic mode. The high selectivity of the approach is illustrated by the dramatically lower interaction with gram-negative bacteria (Escherichia Coli). The double-propulsion (catalytic or fuel-free magnetic) mode of the micromotors and the high biocompatibility holds considerable promise to design micromotors with tailored lanbiotics that can response to the changes that make the bacteria resistant in a myriad of clinical, environmental remediation or food safety applications.
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Antibacterianos/farmacologia , Materiais Biocompatíveis/química , Biofilmes/efeitos dos fármacos , Escherichia coli/fisiologia , Antibacterianos/química , Materiais Biocompatíveis/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Compostos Férricos/química , Grafite/química , Células HeLa , Hemólise/efeitos dos fármacos , Humanos , Luz , Magnetismo , Nanopartículas Metálicas/química , Microscopia de Fluorescência , Nisina/química , Platina/química , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/fisiologia , Imagem com Lapso de TempoRESUMO
Online coupling of a micromotor-based immunoassay and a microfluidic electrochemical detection was explored as a new approach for C-reactive protein (CRP) determination in serum and preterm neonatal plasma samples with sepsis suspicion. The approach combines the advantages of micromotors (self-fluid mixing capabilities leading to a faster assay in low sample volumes) and electrochemical microfluidic (flow-controlled ultraminiaturized electrochemical detection, high sensitivity, and low-cost) technologies. Both technologies elegantly meet the point of care testing or bed side device requirements such as low analysis times, miniaturization and simplification, and single use. Anti-CRP functionalized micromotors (anti-CRP-rGO(reduced graphene oxide)/Ni/PtNPs (platinum nanoparticles))-based immunoassay coupled to thin layer Au-based electrochemical microfluidics operating at -0.20 V under controlled fluidic detection operations (30 µL min-1) allowed the sensitive (LOD = 0.54 µg/mL) and accurate CRP determination using very low volume preterm neonatal clinical samples (<10 µL) in just 8 min of total assay time. These excellent analytical characteristics obtained linked to the full automatization of the immunoassay allowed the fast and accurate determination of CRP in hardly available clinical samples as those coming from preterm infants with suspected sepsis. These results demonstrated the usefulness of the approach which meets the clinical requirements as a future point-of-care device for clinical analysis.
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Técnicas Biossensoriais , Proteína C-Reativa/análise , Técnicas Eletroquímicas , Imunoensaio , Dispositivos Lab-On-A-Chip , Sepse/sangue , Técnicas Biossensoriais/instrumentação , Técnicas Eletroquímicas/instrumentação , Humanos , Imunoensaio/instrumentação , Recém-Nascido , Tamanho da Partícula , Sepse/diagnóstico , Propriedades de SuperfícieRESUMO
WS2/Pt and MoS2/Pt bubble propelled micromotors are used as "on-the-fly" sensing platforms in bioassays, using a highly selective affinity peptide probe for "OFF-ON" detection of Escherichia coli as a model endotoxin. The different outer micromotor surface characteristics play an important role in the sensing performance. The relatively high outer surface of WS2/Pt micromotors results in a 3.5-fold increase in sensitivity compared to MoS2/Pt micromotors due to enhanced peptide probe loading and release. The peptide-modified WS2 micromotors are used as a low-cost and high-throughput approach for the determination of E. coli endotoxin after only 60 s, with a limit of detection of 1.9 ng mL-1 and high selectivity. The method has been validated using spiked samples (tap water, blood serum, and saliva) and bacteria media (blank broth, Staphylococcus aureus, and E. coli). The concept can be extended to the analysis of other (bio)-analytes and easily incorporated into portable instrumentation, holding great promise in a myriad of clinical, environmental, or food-safety applications.