Loading...
50 results
Search Results
Now showing 1 - 10 of 50
- Electrochemistry-assisted surface plasmon resonance detection of miRNA-145 at femtomolar levelPublication . Ribeiro, José A.; Sales, Maria Goreti Ferreira; Pereira, Carlos M.In this work, we combined electrochemical techniques with SPR (eSPR) for the label-free detection of cancer biomarker miRNA-145. Detection was performed in a simple two-step assay. In the first step, the gold sensor surface, previously functionalized with a self-assembled monolayer (SAM) of thiolated RNA probes is incubated with the sample containing the target RNA biomarker. In this step, hybridization of RNA fragments with complementary immobilized probes was monitored in real-time by SPR. In the second step, eSPR measurements were performed to improve the sensitivity of the hybridization assay. Potential-induced deposition of a redox probe at the sensor surface resulted in enhanced SPR response promoted by the electrochemical process, thereby allowing the detection of miRNA-145 at femtomolar level (LOD = 0.56 fM), without sample derivatization or post-hybridization treatment for signal amplification. Good linearity was achieved (R2 = 0.984) over the concentration range from 1.0 fM and 10 nM. Furthermore, the developed eSPR biosensor showed high selectivity towards single-base and two-base mismatch sequences and detection of target miRNA-145 in synthetic human serum was successful achieved.
- An ultrasensitive human cardiac troponin T graphene screen-printed electrode based on electropolymerized-molecularly imprinted conducting polymerPublication . Silva, Bárbara V.M.; Rodríguez, Blanca A.G.; Sales, Goreti; Sotomayor, Maria Del Pilar T.; Dutra, Rosa F.A nano-molecularly imprinted polymer (N-MIP) assembled on a screen-printed electrode for the cardiac troponin T (cTnT) was developed. The biomimetic surface was obtained by a co-polymer matrix as-sembled on the reduced graphene oxide (RGO) electrode surface. The cTnT active sites were engineered using pyrrole and carboxylated pyrrole that was one-step electropolymerized jointly with cTnT by cyclic voltammetry. The stepwise preparation of the biomimetic surface was characterized by cyclic and dif-ferential pulse voltammetries using the ferrocyanide/ferricyanide as redox probe. Structural and mor-phological characterization was also performed. The optimal relation of pyrrole and pyrrole-3-acid car-boxylic to perform the cTnT biomimetic nanosurface was obtained at 1:5 ratio. The analytical perfor-mance of cTnT N-MIP performed by differential pulse voltammetry showed a linear range from 0.01 to 0.1 ng mL-1 (r¼0.995, p«0.01), with a very low limit of detection (0.006 ng mL-1). The synergic effect of conductive polymer and graphene forming 3D structures of reactive sites resulted in a N-MIP with ex-cellent affinity to cTnT binding (KD¼7.3 10-13 mol L-1). The N-MIP proposed is based on a simple method of antibody obtaining with a large potential for point-of-care testing applications.
- Plastic Antibody of Polypyrrole/Multiwall Carbon Nanotubes on Screen-Printed Electrodes for Cystatin C DetectionPublication . Gomes, Rui S.; Gomez-Rodríguez, Blanca Azucena; Fernandes, Ruben; Sales, Goreti; Moreira, Felismina; Dutra, Rosa F.This work reports the design of a novel plastic antibody for cystatin C (Cys-C), an acute kidney injury biomarker, and its application in point-of-care (PoC) testing. The synthetic antibody was obtained by tailoring a molecularly imprinted polymer (MIP) on a carbon screen-printed electrode (SPE). The MIP was obtained by electropolymerizing pyrrole (Py) with carboxylated Py (Py-COOH) in the presence of Cys-C and multiwall carbon nanotubes (MWCNTs). Cys-C was removed from the molecularly imprinted poly(Py) matrix (MPPy) by urea treatment. As a control, a non-imprinted poly(Py) matrix (NPPy) was obtained by the same procedure, but without Cys-C. The assembly of the MIP material was evaluated in situ by Raman spectroscopy and the binding ability of Cys-C was evaluated by the cyclic voltammetry (CV) and differential pulse voltammetry (DPV) electrochemical techniques. The MIP sensor responses were measured by the DPV anodic peaks obtained in the presence of ferro/ferricyanide. The peak currents decreased linearly from 0.5 to 20.0 ng/mL of Cys-C at each 20 min successive incubation and a limit of detection below 0.5 ng/mL was obtained at pH 6.0. The MPPy/SPE was used to analyze Cys-C in spiked serum samples, showing recoveries <3%. This device showed promising features in terms of simplicity, cost and sensitivity for acute kidney injury diagnosis at the point of care.
- Molecular Imprinting on Nanozymes for Sensing ApplicationsPublication . Cardoso, Ana Rita; Frasco, Manuela F.; Serrano, Verónica; Fortunato, Elvira; Sales, Maria Goreti FerreiraAs part of the biomimetic enzyme field, nanomaterial-based artificial enzymes, or nanozymes, have been recognized as highly stable and low-cost alternatives to their natural counterparts. The discovery of enzyme-like activities in nanomaterials triggered a broad range of designs with various composition, size, and shape. An overview of the properties of nanozymes is given, including some examples of enzyme mimics for multiple biosensing approaches. The limitations of nanozymes regarding lack of selectivity and low catalytic efficiency may be surpassed by their easy surface modification, and it is possible to tune specific properties. From this perspective, molecularly imprinted polymers have been successfully combined with nanozymes as biomimetic receptors conferring selectivity and improving catalytic performance. Compelling works on constructing imprinted polymer layers on nanozymes to achieve enhanced catalytic efficiency and selective recognition, requisites for broad implementation in biosensing devices, are reviewed. Multimodal biomimetic enzyme-like biosensing platforms can offer additional advantages concerning responsiveness to different microenvironments and external stimuli. Ultimately, progress in biomimetic imprinted nanozymes may open new horizons in a wide range of biosensing applications.
- An all-in-one approach for self-powered sensing: A methanol fuel cell modified with a molecularly imprinted polymer for cancer biomarker detectionPublication . Carneiro, Liliana P.T; Pinto, Alexandra M.F.R.; Mendes, Adélio; Ferreira Sales, Maria GoretiThis work describes the development of an innovative electrochemical biosensor comprehending a passive direct methanol fuel cell (DMFC) assembly, modified by a layer of a molecularly imprinted polymer (MIP) on a carbon fabric anode electrode containing Pt/Ru nanoparticles. This MIP film was prepared from poly(3,4-ethylenedioxythiophene) (PEDOT) and polypyrrole (PPy) obtained by in situ electropolymerization of the corresponding monomers on the anode electrode surface. This MIP film is designed to detect an important cancer biomarker- carcinoembryonic antigen (CEA). This innovative, all-in-one device works in a simple way. First, CEA is incubated on the anode container of the fuel cell, then methanol is added, followed by the response evaluation (polarization curves determination). As CEA selectively interacts with the MIP film, it blocks the methanol's access to the Pt catalyst, remains specific bonded, and interferes with the subsequent polarization curves of the DMFC. Polarization curves obtained in the presence of standard solutions prepared in buffer and human serum confirmed linear responses of log CEA concentration ranging from 30 to 30 000 ng/mL in both media. The biosensor DMFC showed a sensitive response with a detection limit of 4.41 ng/mL when an aqueous 0.05 M methanol solution was used as fuel. When methanol was replaced by an ethanol solution of the same concentration (using the same setup developed for the DMFC), the lower detection limit of 3.52 ng/mL was obtained. Overall, the obtained results show that methanol/ethanol fuel cells operating without flow-through can be successfully used for the fabrication of self-powered biosensors. The novel biosensor concept presented here is simple, inexpensive, and effective, and can be further developed to meet point-of-care requirements.
- Electrochemical Aptasensor for the Detection of the Key Virulence Factor YadA of Yersinia enterocoliticaPublication . Sande, Maria Georgina; Ferreira, Débora; Rodrigues, Joana; Melo, Luís; Linke, Dirk; Silva, Carla J.; Moreira, Felismina; Sales, Goreti; Rodrigues, LígiaNew point-of-care (POC) diagnosis of bacterial infections are imperative to overcome the deficiencies of conventional methods, such as culture and molecular methods. In this study, we identified new aptamers that bind to the virulence factor Yersinia adhesin A (YadA) of Yersinia enterocolitica using cell-systematic evolution of ligands by exponential enrichment (cell-SELEX). Escherichia coli expressing YadA on the cell surface was used as a target cell. After eight cycles of selection, the final aptamer pool was sequenced by high throughput sequencing using the Illumina Novaseq platform. The sequencing data, analyzed using the Geneious software, was aligned, filtered and demultiplexed to obtain the key nucleotides possibly involved in the target binding. The most promising aptamer candidate, Apt1, bound specifically to YadA with a dissociation constant (Kd) of 11 nM. Apt1 was used to develop a simple electrochemical biosensor with a two-step, label-free design towards the detection of YadA. The sensor surface modifications and its ability to bind successfully and stably to YadA were confirmed by cyclic voltammetry, impedance spectroscopy and square wave voltammetry. The biosensor enabled the detection of YadA in a linear range between 7.0 × 104 and 7.0 × 107 CFU mL−1 and showed a square correlation coefficient >0.99. The standard deviation and the limit of detection was ~2.5% and 7.0 × 104 CFU mL−1, respectively. Overall, the results suggest that this novel biosensor incorporating Apt1 can potentially be used as a sensitive POC detection system to aid the diagnosis of Y. enterocolitica infections. Furthermore, this simple yet innovative approach could be replicated to select aptamers for other (bacterial) targets and to develop the corresponding biosensors for their detection.
- Plastic antibody for the electrochemical detection of bacterial surfaceproteinsPublication . Khan, M. Azizur R.; Moreira, Felismina T. C.; Riu, Jordi; Ferreira Sales, Maria GoretiThis work presents a novel molecularly imprinted polymer (MIP) for the indirect detection of bacteria, by targeting an outer membrane protein on a disposable device. Protein A (PA) was selected for this purpose, as a representative protein of the outer surface of Staphylococcus aureus. The imprinted polymer was assembled directly on a film of single walled carbon nanotubes (SWCNTs), placed on screen-printed electrodes (SPEs). The MIP material was produced by electropolymerizing 3-aminophenol in the presence of the protein template (PA) using cyclic voltammetry (CV). The proteins entrapped at the polymeric backbone were digested by the action of proteolytic activity of proteinase K and then washed away to create vacant sites. The performance of the corresponding imprinted and non-imprinted electrodes was evaluated by EIS and the effect of several variables, such as monomer and template concentrations, or thickness of imprint-ing surface, was controlled and optimized by the number of CV cycles. The detection limit of the MIP-based sensors was 0.60 nM in MES buffer. High repeatability and good selectivity were observed in the presence of a model protein BSA. The sensor performance was also tested to check the effect of inorganic ions in tap water. The detection limit observed was 16.83 nM, with a recovery factor of 91.1 ± 6.6%. The sensor described in this work is a potential tool for screening PA on-site, due to the simplicity of fabrication, disposability, short response time, low cost, good sensitivity and selectivity.
- Screen-printed electrode produced by printed-circuit board technology. Application to cancer biomarker detection by means of plastic antibody as sensing materialPublication . Moreira, Felismina T.C.; Ferreira, M.Judite M.S.; Puga, José R.T.; Sales, GoretiThis research work presents, for the first time, a screen-printed electrode (SPE) made on a PCB board with silver tracks (Ag) and a three electrode configuration (AgxO-working, AgxO-counter and Ag/AgxO-reference electrodes), following the same approach as printed-circuit boards (PCBs). This low cost and disposable device was tested for screening a cancer biomarker in point-of-care. The selected biomarker was carcinogenic embryonic antigen (CEA) protein, routinely used to follow-up the progression of specific cancer diseases. The biosensor was constructed by assembling a plastic antibody on the Ag-working electrode area, acting as the biorecognition element of the device. The protein molecules that were entrapped on the polymer and positioned at the outer surface of the polypyrrole (PPy) film were removed by protease action. The imprinting effect was tested by preparing non-imprinted (NPPy) material, including only PPy as biorecognition element. Infrared and Raman studies confirmed the surface modification of these electrodes. The ability of the sensing material to rebind CEA was measured by several electrochemi-cal techniques: cyclic voltammetry (CV), impedance spectroscopy (EIS) and square wave voltammetry (SWV). The linear response ranged from 0.05 to 1.25 pg/mL against logarithm concentration. Overall, producing screen-printed electrodes by means of conventional PCB technology showed promising features, mostly regarding cost and prompt availability. The plastic antibody-based biosensor also seems to be a promising tool for screening CEA in point-of-care, with low response time, low cost, good sensitivity and high stability.
- Paper-based (bio)sensor for label-free detection of 3-nitrotyrosine in human urine samples using molecular imprinted polymerPublication . Martins, Gabriela V.; Marques, Ana C.; Fortunato, Elvira; Sales, Maria Goreti FerreiraOver the last years, paper technology has been widely spread as a more affordable, sustainable and reliable support material to be incorporated in the design of point-of-care (POC) diagnostic devices. However, the single work employing a paper-based device for 3-nitrotyrosine (3-NT), a relevant biomarker for oxidative stress (OS) that is a major origin for many diseases, is incapable of reading successfully complex samples because every species that oxidizes before ~0.75 V will also contribute to the final response. Thus, the introduction of a selective element was made into this set-up by including a molecularly-imprinted polymer (MIP) tailored in-situ. Herein, a novel MIP for 3-NT was assembled directly on a paper platform, made conductive with carbon ink and suitable for an electrochemical transduction. The biomimetic material was produced by electropolymerization of phenol after optimizing several experimental parameters, such a scan-rate, number of cycles, range of potential applied, monomer and template concentrations. Under optimal conditions, the label-free sensor was able to respond to 3-NT from 500 nM to 1 mM, yielding a limit of detection of 22.3 nM. Finally, the applicability of the (bio)sensor was tested by performing calibration assays in human urine samples and a good performance was obtained in terms of sensitivity, selectivity and reproducibility. Overall, the attributes of the herein described sensing approach can be compared to a very limited number of other electrochemical devices, that are still using a conventional three electrode system, making this paper-sustained device the first electrochemical (bio)sensor with potential to become a portable and low-cost diagnostic tool for 3-NT. In general, the incorporation of molecular imprinting technology coupled to electrochemical transduction enabled the fabrication of suitable smart sensors for wide screening approaches.
- Photonics in nature and bioinspired designs: sustainable approaches for a colourful worldPublication . Vaz, Raquel; Frasco, Manuela F.; Sales, M. Goreti F.Biological systems possess nanoarchitectures that have evolved for specific purposes and whose ability to modulate the flow of light creates an extraordinary diversity of natural photonic structures. In particular, the striking beauty of the structural colouration observed in nature has inspired technological innovation in many fields. Intense research has been devoted to mimicking the unique vivid colours with newly designed photonic structures presenting stimuli-responsive properties, with remarkable applications in health care, safety and security. This review highlights bioinspired photonic approaches in this context, starting by presenting many appealing examples of structural colours in nature, followed by describing the versatility of fabrication methods and designed coloured structures. A particular focus is given to optical sensing for medical diagnosis, food control and environmental monitoring, which has experienced a significant growth, especially considering the advances in obtaining inexpensive miniaturized systems, more reliability, fast responses, and the use of label-free layouts. Additionally, naturally derived biomaterials and synthetic polymers are versatile and fit many different structural designs that are underlined. Progress in bioinspired photonic polymers and their integration in novel devices is discussed since recent developments have emerged to lift the expectations of smart, flexible, wearable and portable sensors. The discussion is expanded to give emphasis on additional functionalities offered to related biomedical applications and the use of structural colours in new sustainable strategies that could meet the needs of technological development.