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Lead ions are especially harmful to human health, causing significant developmental and behavioral abnormalities even at small concentrations. In real-life samples, lead ions are present in mixtures with other metal ions, creating a challenge to detect it selectively at low quantities. To address these challenges, we prepared an electrochemical sensor based on delaminated Ti3C2Tx MXene, which can selectively detect low concentrations of Pb2+ in a solution containing other common metal ions. Cyclic voltammetry was applied as an electrochemical detection method. The proposed reaction mechanism involves a reversible transition between Pb2+ ions and PbO at the MXene-based layer. The sensitivity of the sensor towards Pb2+ ions and a limit of detection were determined. The sensor, as prepared, had a linear response range within 0.15-1.0 µM, with a sensitivity of 26.7 µA/µM and LOD value of 48.7 nM, which meets the requirements set by the World Health Organization.
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This study addresses the need for enhanced antimicrobial properties of electrospun membranes, either through surface modifications or the incorporation of antimicrobial agents, which are crucial for improved clinical outcomes. In this context, chitosan-a biopolymer lauded for its biocompatibility and extracellular matrix-mimicking properties-emerges as an excellent candidate for tissue regeneration. However, fabricating chitosan nanofibers via electrospinning often challenges the preservation of their structural integrity. This research innovatively develops a chitosan/polycaprolactone (CH/PCL) composite nanofibrous membrane by employing a layer-by-layer electrospinning technique, enhanced with silver nanoparticles (AgNPs) synthesized through a wet chemical process. The antibacterial efficacy, adhesive properties, and cytotoxicity of electrospun chitosan membranes were evaluated, while also analyzing their hydrophilicity and nanofibrous structure using SEM. The resulting CH/PCL-AgNPs composite membranes retain a porous framework, achieve balanced hydrophilicity, display commendable biocompatibility, and exert broad-spectrum antibacterial activity against both Gram-negative and Gram-positive bacteria, with their efficacy correlating to the AgNP concentration. Furthermore, our data suggest that the antimicrobial efficiency of these membranes is influenced by the timed release of silver ions during the incubation period. Membranes incorporated starting with AgNPs at a concentration of 50 µg/mL effectively suppressed the growth of both microorganisms during the early stages up to 8 h of incubation. These insights underscore the potential of the developed electrospun composite membranes, with their superior antibacterial qualities, to serve as innovative solutions in the field of tissue engineering.
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This paper reports on development of an optical biosensor for the detection of antibodies against SARS-CoV-2 virus proteins in blood serum. ZnO nanotetrapods with high surface area and stable room temperature photoluminescence (PL) were selected as transducers. Structure and optical properties of the ZnO tetrapods have been studied by XRD, SEM and Raman spectroscopy. Crystallinity, dimensions and emission peaks of the ZnO tetrapods were determined. The ZnO tetrapods were fixed on glass chip. Silanization of ZnO tetrapods surface resulted in forming of functional surface groups suitable for the immobilization of bioselective layer. Two types of recombinant proteins (rS and rN) have been used to form bioselective layer on the surface of the ZnO tetrapods. Flow through microfluidic system, integrated with optical system, has been used for the determination of antibodies against SARS-CoV-2 virus proteins present in blood samples. The SARS-CoV-2 probes, prepared in PBS solution, have been injected into the measurement chamber with a constant pumping speed. Steady-state photoluminescence spectra and photoluminescence kinetics have been studied before and after injection of the probes. The biosensor signal has been tested to anti-SARS-CoV-2 antibodies in the range of 0.001 nM-1 nM. Control measurements have been performed with blood serum of healthy person. ZnO-SARS-CoV-2-rS and ZnO-SARS-CoV-2-rN biosensors showed high stability and sensitivity to anti-SARS-CoV-2 antibodies in the range of 0.025-0.5 nM (LOD 0.01 nM) and 0.3-1 nM (LOD 0.3 nM), respectively. Gibbs free energy of interaction between ZnO/SARS-CoV-2-rS and ZnO/SARS-CoV-2-rN bioselective layers with anti-SARS-CoV-2 antibodies showed -35.5 and -21.4 kJ/mol, respectively. Average detection time of biosensor integrated within microfluidic system was 15-20 min. The detection time and pumping speed (50 µL/min) were optimized to make detection faster. The developed system and ZnO-SARS-CoV-2-rS nanostructures have good potential for detection of anti-SARS-CoV-2 antibodies from patient's probes.
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Anticorpos Antivirais , Técnicas Biossensoriais , SARS-CoV-2 , Óxido de Zinco , Óxido de Zinco/química , Técnicas Biossensoriais/instrumentação , Técnicas Biossensoriais/métodos , SARS-CoV-2/imunologia , Anticorpos Antivirais/sangue , Humanos , COVID-19 , Medições Luminescentes/métodos , Microfluídica/métodosRESUMO
Extending applications of Ti3C2Tx MXene in nanocomposites and across fields of electronics, energy storage, energy conversion, and sensor technologies necessitates simple and efficient analytical methods. Raman spectroscopy is a critical tool for assessing MXene composites; however, high laser powers and temperatures can lead to the materials' deterioration during the analysis. Therefore, an in-depth understanding of MXene photothermal degradation and changes in its oxidation state is required, but no systematic studies have been reported. The primary aim of this study was to investigate the degradation of the MXene lattice through Raman spectroscopic analysis. Distinct spectral markers were related to structural alterations within the Ti3C2Tx material after subjecting it to thermal- and laser-induced degradation. During the degradation processes, spectral markers were revealed for several specific steps: a decrease in the number of interlayer water molecules, a decrease in the number of -OH groups, formation of C-C bonds, oxidation of the lattice, and formation of TiO2 nanoparticles (first anatase, followed by rutile). By tracking of position shifts and intensity changes for Ti3C2Tx, the spectral markers that signify the initiation of each step were found. This spectroscopic approach enhances our understanding of the degradation pathways of MXene, and facilitating enhanced and dependable integration of these materials into devices for diverse applications, from energy storage to sensors.
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A living cell is a complex network of molecular, biochemical and physiological processes. Cellular activities, such as ion transport, metabolic processes, and cell-cell interactions can be determined electrochemically by detecting the electrons or ions exchanged in these processes. Electrochemical methods often are noninvasive, and they can enable the real-time monitoring of cellular processes. Scanning electrochemical microscopy (SECM) is an advanced scanning probe electroanalysis technique that can map the surface topography and local reactivity of a substrate with high precision at the micro- or nanoscale. By measuring electrochemical signals, such as redox reactions, ion fluxes, and pH changes, SECM can provide valuable insights into cellular activity. As a result of its compatibility with liquid medium measurements and its nondestructive nature, SECM has gained popularity in living cell research. This review aims to furnish an overview of SECM, elucidating its principles, applications, and its potential to contribute significantly to advancements in cell biology, electroporation, and biosensors. As a multidisciplinary tool, SECM is distinguished by its ability to unravel the intricacies of living cells and offers promising avenues for breakthroughs in our understanding of cellular complexity.
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MXenes have received worldwide attention across various scientific and technological fields since the first report of the synthesis of Ti3C2 nanostructures in 2011. The unique characteristics of MXenes, such as superior mechanical strength and flexibility, liquid-phase processability, tunable surface functionality, high electrical conductivity, and the ability to customize their properties, have led to the widespread development and exploration of their applications in energy storage, electronics, biomedicine, catalysis, and environmental technologies. The significant growth in publications related to MXenes over the past decade highlights the extensive research interest in this material. One area that has a great potential for improvement through the integration of MXenes is sensor design. Strain sensors, temperature sensors, pressure sensors, biosensors (both optical and electrochemical), gas sensors, and environmental pollution sensors targeted at volatile organic compounds (VOCs) could all gain numerous improvements from the inclusion of MXenes. This report delves into the current research landscape, exploring the advancements in MXene-based chemo-sensor technologies and examining potential future applications across diverse sensor types.
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In this study, we demonstrate that palladium-platinum bimetallic nanoparticles (Pd@Pt NPs) as the nanozyme, combined with a multi-layer paper-based analytical device and DNA hybridization, can successfully detect Mycobacterium tuberculosis. This nanozyme has peroxidase-like properties, which can increase the oxidation rate of the substrate. Compared with horseradish peroxidase, which is widely used in traditional detection, the Michaelis constants of Pd@Pt NPs are fourteen and seventeen times lower than those for 3,3',5,5'-tetramethylbenzidine and H2O2, respectively. To verify the catalytic efficiency of Pd@Pt NPs, this study will execute molecular diagnosis of Mycobacterium tuberculosis. We chose the IS6110 fragment as the target DNA and divided the complementary sequences into the capture DNA and reporter DNA. They were modified on paper and Pd@Pt NPs, respectively, to detect Mycobacterium tuberculosis on a paper-based analytical device. With the above-mentioned method, we can detect target DNA within 15 minutes with a linear range between 0.75 and 10 nM, and a detection limit of 0.216 nM. These results demonstrate that the proposed platform (a DNA-nanozyme integrated paper-based analytical device, dnPAD) can provide sensitive and on-site infection prognosis in areas with insufficient medical resources.
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Nanopartículas Metálicas , Mycobacterium tuberculosis , Peróxido de Hidrogênio/química , Platina/química , Paládio/química , Nanopartículas Metálicas/química , DNA , ColorimetriaRESUMO
This research is focused on enhancing the capabilities of scanning electrochemical impedance microscopy (SEIM) for detecting gold nanoparticle-labelled antibodies using electrochemically modified platinum ultramicroelectrode. The primary objective was to address the high resistance issue encountered in previous measurements with SEIM via the utilization of SEIM probes based on micro-electrodes modified by platinum microstructures, which improved the sensitivity and precision of the detection of targeted biomolecules. The modified probe resulted in a lowered charge transfer resistance by over ten times and a decrease in detection to around 100 fg/mL. We suggest potential applications in various biotechnological and biomedical fields, with future research expected to further refine this technique.
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The emergence of COVID-19 caused by the coronavirus SARS-CoV-2 has prompted a global pandemic that requires continuous research and monitoring. This study presents a design of an electrochemical biosensing platform suitable for the evaluation of monoclonal antibodies targeting the SARS-CoV-2 nucleocapsid (N) protein. Screen-printed carbon electrodes (SPCE) modified with gold nanostructures (AuNS) were applied to design a versatile and sensitive sensing platform. Electrochemical techniques, including electrochemical impedance spectroscopy (EIS) and square wave voltammetry (SWV), were used to investigate the interactions between immobilised recombinant N (rN) protein and several monoclonal antibodies (mAbs). The electrochemical characterisation of SPCE/AuNS/rN demonstrated a successful immobilisation of rN, enhancing the electron transfer kinetics. Affinity interactions between immobilised rN and four mAbs (mAb-4B3, mAb-4G6, mAb-12B2, and mAb-1G5) were explored. Although mAb-4B3 showed some non-linearity, the other monoclonal antibodies exhibited specific and well-defined interactions followed by the formation of an immune complex. The biosensing platform demonstrated high sensitivity in the linear range (LR) from 0.2 nM to 1 nM with limits of detection (LOD) ranging from 0.012 nM to 0.016 nM for mAb-4G6, mAb-12B2, and mAb-1G5 and limits of quantification (LOQ) values ranging from 0.035 nM to 0.139 nM, as determined by both EIS and SWV methods. These results highlight the system's potential for precise and selective detection of monoclonal antibodies specific to the rN. This electrochemical biosensing platform provides a promising route for the sensitive and accurate detection of monoclonal antibodies specific to the rN protein.
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Técnicas Biossensoriais , COVID-19 , Humanos , SARS-CoV-2 , Anticorpos Monoclonais , Limite de Detecção , Técnicas Eletroquímicas/métodos , Carbono , Técnicas Biossensoriais/métodos , EletrodosRESUMO
This article aims to present a comparative study of three polypyrrole-based molecularly imprinted polymer (MIP) systems for the detection of the recombinant severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein (rN). The rN is known for its relatively low propensity to mutate compared to other SARS-CoV-2 antigens. The aforementioned systems include screen-printed carbon electrodes (SPCE) modified with gold nanostructures (MIP1), platinum nanostructures (MIP2), and the unmodified SPCE (MIP3), which was used for control. Pulsed amperometric detection (PAD) was employed as the detection technique, offering the advantage of label-free detection without the need for an additional redox probe. Calibration curves were constructed using the obtained data to evaluate the response of each system. Non-imprinted systems were also tested in parallel to evaluate the contribution of non-specific binding and assess the affinity sensor's efficiency. The analysis of calibration curves revealed that the AuNS-based MIP1 system exhibited the lowest contribution of non-specific binding and displayed a better fit with the chosen fitting model compared to the other systems. Further analysis of this system included determining the limit of detection (LOD) (51.2 ± 2.8 pg/mL), the limit of quantification (LOQ) (153.9 ± 8.3 pg/mL), and a specificity test using a recombinant receptor-binding domain of SARS-CoV-2 spike protein as a control. Based on the results, the AuNS-based MIP1 system demonstrated high specificity and sensitivity for the label-free detection of SARS-CoV-2 nucleocapsid protein. The utilization of PAD without the need for additional redox probes makes this sensing system convenient and valuable for rapid and accurate virus detection.
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Técnicas Biossensoriais , COVID-19 , Glicoproteína da Espícula de Coronavírus , Humanos , SARS-CoV-2 , COVID-19/diagnóstico , Polímeros/química , Pirróis , Proteínas do Nucleocapsídeo/análiseRESUMO
The emergence of antibiotic-resistant bacteria, particularly the most hazardous pathogens, namely Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp. (ESKAPE)-pathogens pose a significant threat to global health. Current antimicrobial therapies, including those targeting biofilms, have shown limited effectiveness against these superbugs. Nanoparticles, specifically silver nanoparticles (AgNPs), have emerged as a promising alternative for combating bacterial infections. In this study, two types of AgNPs with different physic-chemical properties were evaluated for their antimicrobial and antibiofilm activities against clinical ESKAPE strains. Two types of silver nanoparticles were assessed: spherical silver nanoparticles (AgNPs-1) and cubic-shaped silver nanoparticles (AgNPs-2). AgNPs-2, characterized by a cubic shape and higher surface-area-to-volume ratio, exhibited superior antimicrobial activity compared to spherical AgNPs-1. Both types of AgNPs demonstrated the ability to inhibit biofilm formation and disrupt established biofilms, leading to membrane damage and reduced viability of the bacteria. These findings highlight the potential of AgNPs as effective antibacterial agents against ESKAPE pathogens, emphasizing the importance of nanoparticle characteristics in determining their antimicrobial properties. Further research is warranted to explore the underlying mechanisms and optimize nanoparticle-based therapies for the management of infections caused by antibiotic-resistant bacteria.
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Molecularly imprinted polymers (MIPs), a type of biomimetic material, have attracted considerable interest owing to their cost-effectiveness, good physiochemical stability, favourable specificity and selectivity for target analytes, and widely used for various biological applications. It was demonstrated that MIPs with significant selectivity towards protein-based targets could be applied in medicine, diagnostics, proteomics, environmental analysis, sensors, various in vivo and/or in vitro applications, drug delivery systems, etc. This review provides an overview of MIPs dedicated to biomedical applications and insights into perspectives on the application of MIPs in newly emerging areas of biotechnology. Many different protocols applied for the synthesis of MIPs are overviewed in this review. The templates used for molecular imprinting vary from the minor glycosylated glycan-based structures, amino acids, and proteins to whole bacteria, which are also overviewed in this review. Economic, environmental, rapid preparation, stability, and reproducibility have been highlighted as significant advantages of MIPs. Particularly, some specialized MIPs, in addition to molecular recognition properties, can have high catalytic activity, which in some cases could be compared with other bio-catalytic systems. Therefore, such MIPs belong to the class of so-called 'artificial enzymes'. The discussion provided in this manuscript furnishes a comparative analysis of different approaches developed, underlining their relative advantages and disadvantages highlighting trends and possible future directions of MIP technology.
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Impressão Molecular , Impressão Molecular/métodos , Reprodutibilidade dos Testes , Polímeros/química , Proteínas , Sistemas de Liberação de MedicamentosRESUMO
Recently, rare diseases have received attention due to the need for improvement in diagnosed patients' and their families' lives. Duchenne muscular dystrophy (DMD) is a rare, severe, progressive, muscle-wasting disease. Today, the therapeutic standard for treating DMD is corticosteroids, which cause serious adverse side effects. Nutraceuticals, e.g., herbal extracts or essential oils (EOs), are possible active substances to develop new drug delivery systems to improve DMD patients' lives. New drug delivery systems lead to new drug effects, improved safety and accuracy, and new therapies for rare diseases. Herbal extracts and EOs combined with click chemistry can lead to the development of safer treatments for DMD. In this review, we focus on the need for novel drug delivery systems using EOs as the therapy for DMD and the potential use of click chemistry for drug delivery systems. New EO complex drug delivery systems may offer a new approach for improving muscle conditions and mental health issues associated with DMD. However, further research should identify the potential of these systems in the context of DMD. In this review, we discuss possibilities for applying EOs to DMD before implementing expensive research in a theoretical way.
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Synthetic bone grafting materials play a significant role in various medical applications involving bone regeneration and repair. Their ability to mimic the properties of natural bone and promote the healing process has contributed to their growing relevance. While calcium-phosphates and their composites with various polymers and biopolymers are widely used in clinical and experimental research, the diverse range of available polymer-based materials poses challenges in selecting the most suitable grafts for successful bone repair. This review aims to address the fundamental issues of bone biology and regeneration while providing a clear perspective on the principles guiding the development of synthetic materials. In this study, we delve into the basic principles underlying the creation of synthetic bone composites and explore the mechanisms of formation for biologically important complexes and structures associated with the various constituent parts of these materials. Additionally, we offer comprehensive information on the application of biologically active substances to enhance the properties and bioactivity of synthetic bone grafting materials. By presenting these insights, our review enables a deeper understanding of the regeneration processes facilitated by the application of synthetic bone composites.
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The profound understanding and detailed evaluation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (SCoV2-S) protein and specific antibody interaction mechanism is of high importance in the development of immunosensors for COVID-19. In the present work, we studied a model system of immobilized SCoV2-S protein and specific monoclonal antibodies by molecular dynamics of immune complex formation in real time. We simultaneously applied spectroscopic ellipsometry and quartz crystal microbalance with dissipation to reveal the features and steps of the immune complex formation. We showed direct experimental evidence based on acoustic and optical measurements that the immune complex between covalently immobilized SCoV2-S and specific monoclonal antibodies is formed in two stages. Based on these findings it was demonstrated that applying a two-step binding mathematical model for kinetics analysis leads to a more precise determination of interaction rate constants than that determined by the 1:1 Langmuir binding model. Our investigation showed that the equilibrium dissociation constants (KD) determined by a two-step binding model and the 1:1 Langmuir model could differ significantly. The reported findings can facilitate a deeper understanding of antigen-antibody immune complex formation steps and can open a new way for the evaluation of antibody affinity towards corresponding antigens.
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Técnicas Biossensoriais , COVID-19 , Humanos , Glicoproteína da Espícula de Coronavírus , Complexo Antígeno-Anticorpo , Afinidade de Anticorpos , Imunoensaio , SARS-CoV-2 , Anticorpos MonoclonaisRESUMO
In this study, we are reporting a novel electrochemical capacitance spectroscopy (ECS) platform designed for the sensitive and label-free detection of antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus spike protein (anti-rS) in diluted blood serum. The determination of anti-rS is crucial for identification individuals who have been infected by SARS-CoV-2 virus and may have acquired immunity. The rS protein was immobilized on a screen-printed carbon electrode, which was incubated in diluted blood serum containing anti-rS antibodies. Label-free ECS was applied for the determination of interaction between immobilized rS and free-standing anti-rS. Here reported bioanalytical platform demonstrated high sensitivity and specificity in detecting anti-rS, achieving a limit of detection of 4.38 nM. This versatile platform could be further enhanced by applying various electrode materials and adapting this platform to detect antibodies against some other proteins. Our findings have significant implications for the development of affordable, scalable biosensing platforms capable to provide rapid and accurate public health screening and monitoring, particularly in the context of the coronavirus disease 2019 (COVID-19) pandemic.
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The spike (S) protein and its receptor-binding domain (RBD) of the coronavirus SARS-CoV-2 have been continually evolving, yielding the majority of significant missense mutations and new variants of concern. In this study, we examined how monoclonal antibodies against RBD (mAbs-SCoV2-RBD) and polyclonal antibodies present in convalescent human serum specifically interact with the S protein of wild-type and SARS-CoV-2 variants of concern (VOCs) in real time and how this can be reflected through surface mass density. Moreover, we combined two distinct, label-free measurement techniques: one based on changes in surface electromagnetic waves after reflection from the surface, and the other on changes in acoustic waves. The results demonstrated that dry surface mass density (ΓSE) of mAbs-SCoV2-RBD attached to the RBD of the S protein decreases three-fold, from 148 ng/cm2 to 46 ng/cm2, due to the B.1.351 or so-called beta mutation of coronavirus and its S protein (SCoV2-ß). Consequently, the obtained wet mass ΓQCM-D resulted in values two times lower, from 319 ng/cm2 to 158 ng/cm2, and the hydration of mAbs-SCoV2-RBD/SCoV2-ß immune complex was 70.88%. Conversely, when polyclonal antibodies present in convalescent human serum form immune complexes with the S protein of SARS-CoV-2 variants of concern, the ΓSE decreased from 279 ng/cm2 to 249 ng/cm2, and ΓQCM-D from 1545 ng/cm2 to 1366 ng/cm2. These results can give insights into the differences between the interaction of monoclonal and polyclonal antibodies with SARS-CoV-2 VOCs.
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Anticorpos Monoclonais , COVID-19 , Humanos , SARS-CoV-2 , Glicoproteína da Espícula de CoronavírusRESUMO
The appearance of biological molecules, so-called biomarkers in body fluids at abnormal concentrations, is considered a good tool for detecting disease. Biomarkers are usually looked for in the most common body fluids, such as blood, nasopharyngeal fluids, urine, tears, sweat, etc. Even with significant advances in diagnostic technology, many patients with suspected infections receive empiric antimicrobial therapy rather than appropriate treatment, which is driven by rapid identification of the infectious agent, leading to increased antimicrobial resistance. To positively impact healthcare, new tests are needed that are pathogen-specific, easy to use, and produce results quickly. Molecularly imprinted polymer (MIP)-based biosensors can achieve these general goals and have enormous potential for disease detection. This article aimed to overview recent articles dedicated to electrochemical sensors modified with MIP to detect protein-based biomarkers of certain infectious diseases in human beings, particularly the biomarkers of infectious diseases, such as HIV-1, COVID-19, Dengue virus, and others. Some biomarkers, such as C-reactive protein (CRP) found in blood tests, are not specific for a particular disease but are used to identify any inflammation process in the body and are also under consideration in this review. Other biomarkers are specific to a particular disease, e.g., SARS-CoV-2-S spike glycoprotein. This article analyzes the development of electrochemical sensors using molecular imprinting technology and the used materials' influence. The research methods, the application of different electrodes, the influence of the polymers, and the established detection limits are reviewed and compared.
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Anti-Infecciosos , COVID-19 , Doenças Transmissíveis , Impressão Molecular , Humanos , Polímeros Molecularmente Impressos , Técnicas Eletroquímicas/métodos , SARS-CoV-2 , Doenças Transmissíveis/diagnóstico , Impressão Molecular/métodos , Biomarcadores , Proteína C-Reativa , Eletrodos , Limite de Detecção , Teste para COVID-19RESUMO
Maxillary sinus augmentation is a commonly used procedure for the placement of dental implants. However, the use of natural and synthetic materials in this procedure has resulted in postoperative complications ranging from 12% to 38%. To address this issue, we developed a novel calcium deficient HA/ß-TCP bone grafting nanomaterial using a two-step synthesis method with appropriate structural and chemical parameters for sinus lifting applications. We demonstrated that our nanomaterial exhibits high biocompatibility, enhances cell proliferation, and stimulates collagen expression. Furthermore, the degradation of ß-TCP in our nanomaterial promotes blood clot formation, which supports cell aggregation and new bone growth. In a clinical trial involving eight cases, we observed the formation of compact bone tissue 8 months after the operation, allowing for the successful installation of dental implants without any early postoperative complications. Our results suggest that our novel bone grafting nanomaterial has the potential to improve the success rate of maxillary sinus augmentation procedures.
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We propose a simple, fast, and low-cost method for producing Au-coated black Si-based SERS-active substrates with a proven enhancement factor of 106. Room temperature reactive ion etching of silicon wafer followed by nanometer-thin gold sputtering allows the formation of a highly developed lace-type Si surface covered with homogeneously distributed gold islands. The mosaic structure of deposited gold allows the use of Au-uncovered Si domains for Raman peak intensity normalization. The fabricated SERS substrates have prominent uniformity (with less than 6% SERS signal variations over large areas, 100 × 100 µm2). It has been found that the storage of SERS-active substrates in an ambient environment reduces the SERS signal by less than 3% in 1 month and not more than 40% in 20 months. We showed that Au-coated black Si-based SERS-active substrates can be reused after oxygen plasma cleaning and developed relevant protocols for removing covalently bonded and electrostatically attached molecules. Experiments revealed that the Raman signal of 4-MBA molecules covalently bonded to the Au coating measured after the 10th cycle was just 4 times lower than that observed for the virgin substrate. A case study of the reusability of the black Si-based substrate was conducted for the subsequent detection of 10-5 M doxorubicin, a widely used anticancer drug, after the reuse cycle. The obtained SERS spectra of doxorubicin were highly reproducible. We demonstrated that the fabricated substrate permits not only qualitative but also quantitative monitoring of analytes and is suitable for the determination of concentrations of doxorubicin in the range of 10-9-10-4 M. Reusable, stable, reliable, durable, low-cost Au-coated black Si-based SERS-active substrates are promising tools for routine laboratory research in different areas of science and healthcare.