Browsing by Author "Sales, Maria Goreti Ferreira"
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- A biomimetic photonic crystal sensor for label-free detection of urinary venous thromboembolism biomarkerPublication . Resende, Sara; Frasco, Manuela F.; Sales, Maria Goreti FerreiraThis work reports a novel label-free sensor combining photonic crystals and molecularly-imprinted polymer materials for targeting a protein in human samples within levels of clinical interest. This concept was applied to detect fibrinopeptide B (FPB) in urine, a biomarker of venous thromboembolism, which is a disease of great concern and demands innovative point-of-care devices for improved diagnosis. The molecularly-imprinted photonic polymer (MIPP) was obtained by tailoring an imprinted polymer on highly ordered silica nanoparticles assembled by vertical deposition. Owing to the hierarchical structure, the resulting MIPP exhibited optical properties that changed upon rebinding of the target analyte, FPB. Thus, the changes in reflectance intensity enabled a rapid and sensitive detection of FPB in human urine. The observed response was linear between 0.2 ng mL−1 and 22 ng mL-1 and showed a limit of detection of 0.13 ng mL-1. These features of the sensing material allow the assessment of urinary FPB at relevant clinical levels. In addition, the sensor was selective for FPB compared to the standard biomarker of venous thromboembolism, D-Dimer. The stability of the material was evidenced by the reusability study, which demonstrated reversibility of the maximum intensity after three cycles of recognition and regeneration. Moreover, the sensor showed good performance for the spiked FPB detection in control human urine. Overall, the application of such label-free sensor offers high selectivity and stability, as well as easy operation. It may constitute an alternative method for non-invasive and real-time detection of various protein biomarkers in point-of-care.
- A cellulose-based colour test-strip for equipment-free drug detection on-site: application to sulfadiazine in aquatic environmentPublication . Teixeira, Carla N. O.; Sales, Maria Goreti FerreiraThis work develops a simple and innovative test-strip to monitor antibiotics in aquaculture facilities by an equipment-free approach. It consists of a low-cost disposable cellulose paper that was chemically modified to produce a colour change when in contact with a given antibiotic. In brief, the cellulose substrate was subject to oxidation with periodate, followed by amination with chitosan binding and modification with Cu(II). The test strip was then dipped in the target solution and the intensity of the colour generated therein revealed the concentration of antibiotic present for concentrations higher than 0.5 mM. The higher the concentration in sulfadiazine (SDZ), the more intense the pink colour formed in the final solution, which was also turbid due to the insolubility of the formed product. This colour intensity also varied linearly with the logarithm of the SDZ concentration (from 0.5 to 5 mM), when plotted against the sum of the RGB coordinates extracted from digital pictures. The linear equation of this response was represented by (R + G + B) = − 256.1 log(SDZ, mol/L) − 362.0, with an R-squared of 0.9913. The test-strip was stable for at least 15 days and was selective in the presence of tetracycline and difloxacin, while the response to other members of the sulfadiazine family requires prior evaluation. Overall, the test-strips developed herein are inexpensive and provide valuable (semi-) quantitative data for monitoring SDZ in waters, a most valuable approach to control and reduce the level of antibiotics in fish tanks, which in turn may reduce the costs of fish production and the environmental concerns linked to this practice. Moreover, the test strip uses a cellulose substrate that has little environmental impact upon discard.
- Cellulose‑based hydrogel on quantum dots with molecularly imprinted polymers for the detection of CA19‑9 protein cancer biomarkerPublication . Piloto, Ana Margarida L.; Ribeiro, David S.M.; Rodrigues, S. Sofia M.; Santos, João L.M.; Sampaio, Paula; Sales, Maria Goreti FerreiraMolecularly imprinted polymers (MIPs), were successfully assembled around quantum dots (QDs), for the detection of the carbohydrate antigen CA19-9, a biomarker associated with pancreatic cancer (PC). The imprinted materials MIP@QDs were incorporated within cellulose hydrogels and designated as imprinted hydrogels (iHGs). The concept is to use MIPs which function as the biorecognition elements, conjugated to QDs as the sensing system. The fluorescence quenching of the iHGs occurred with increasing concentrations of CA19-9, showing linearity in the range 2.76 × 10−2 – 5.23 × 102 U/ml, in a 1000-fold diluted human serum. The iHGs show a linear response below the cutoff values for pancreatic cancer diagnosis (< 23 U/ml), a limit of detection of 1.58 × 10−3 U/ml and an imprinting factor (IF) of 1.76. Moreover, they exhibit stability and selectivity towards CA19-9, when compared with the non-imprinted controls, here designated as non-imprinted hydrogels (non-iHGs). In conclusion, this work demonstrates that the conjugation of MIPs to QDs increases the sensitivity and specificity of the developed sensing system for optical detection methods within clinical significance ranges. This fact shows potential for the iHGs to be applied as a sensitive, low-cost format for point-of-care tests (PoCTs).
- Development of an electrochemical biosensor for Galectin-3 detection in point-of-carePublication . Cerqueira, Sofia M.V.; Fernandes, Rúben; Moreira, Felismina; Sales, Maria Goreti FerreiraThis research work aims the development and optimization of an electrochemical biosensor based on molecularly-imprinted polymers [MIPs], for monitoring a melanoma biomarker, Galectin-3 (Gal-3). As it is a multifunctional protein that plays an important role in different types of tumors including melanoma, and has shown good results as a potential biomarker in several areas, the construction of a biosensor for the detection of this protein would be a simple and quick strategy to support the treatment of this type of pathology. The target molecule was recognized by a MIP material, created on the electrode’s surface by electropolymerizing a mixture of analyte (Gal-3) and monomer (2-aminophenol). Then, the protein was removed from the polymeric material by oxalic acid treatment. This process formed a non-conductive polymer with recognition sites showing affinity for the Gal-3. The control of the surface modification was monitored by Raman spectroscopy and electroanalytical techniques, namely electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). The analytical performance of the sensor was evaluated by EIS, by following the analytical response of standard solutions ranging from 0.5 ng/mL to 5000 ng/mL Gal-3 in spyked serum. In general, the biosensor displayed good analytical features, considering limit of detection, response time and reproducibility. Overall, this study resulted from the need to create a new strategy for monitoring melanoma through the creation of a cheaper, faster and sensitive device, which can be commercialized and thus integrate the entire process associated with the treatment and follow-up of this pathology.
- Electrochemical Point-of Care (PoC) Determination of Interleukin-6 (IL-6) Using a Pyrrole (Py) Molecularly Imprinted Polymer (MIP) on a Carbon-Screen Printed Electrode (C-SPE)Publication . Gonçalves, M. de Lurdes; Truta, Liliana; Sales, Maria Goreti Ferreira; Moreira, FelisminaHere is presented a highly sensitive biomimetic sensor for the detection of an Alzheimer’s Disease (AD) biomarker, interleukin-6 (IL6), for point-of-care (PoC) analysis. The imprinted polymeric film was prepared by the co-electropolymerization of pyrrole (Py) and carboxylated pyrrole in the presence of IL6, on a carbon-screen printed electrode . The biomarker molecule was then removed by oxalic acid , creating the recognition vacant sites. Similarly, a control was also prepared in absense of the IL6 biomarker . The different steps of the sensor fabrication were characterized by Raman Spectroscopy, Electrochemical Impedance Spectroscopy (EIS), and Cyclic Voltammetry. Biomarker recognition and capture capacity of the sensing material was measured by EIS. The biomimetic sensor showed a limit of detection for IL6 of 0.02 pg/mL in spiked serum samples. Overall, the biosensing device showed good sensitivity, reproducibility, accuracy, and rapid response time, which contibute for the development of early diagnostics Poc tools for neurological diseases.
- 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.
- Employing bacteria machinery for antibiotic detection: Using DNA gyrase for ciprofloxacin detectionPublication . Cardoso, Ana Rita; Carneiro, Liliana P.T.; Cabral-Miranda, Gustavo; Bachmann, Martin F.; Sales, Maria Goreti FerreiraThis work describes a new successful approach for designing biosensors that detect antibiotics. It makes use of a biomimetic strategy, by employing the biochemical target of a given antibiotic as its biorecognition element. This principle was tested herein for quinolones, which target DNA gyrase in bacteria. Ciprofloxacin (CIPRO) was tested as a representative antibiotic from the quinolone group; the sensitivity of biosensor to this group was confirmed by checking the response to another quinolone antibiotic (norfloxacin, NOR) and to a non-quinolone antibiotic (ampicillin, AMP). The biorecognition element used was DNA gyrase attached by ionic interactions to a carbon support, on a working electrode on common screen-printed electrodes (SPEs). The response against antibiotics was tested for increasing concentrations of CIPRO, NOR or AMP, and following the subsequent electrical changes by electrochemical impedance spectroscopy. The DNAgyrase biosensor showed sensitive responses for CIPRO and NOR, for concentrations down to 3.02 nM and 30.2 nM, respectively, with a very wide response range for CRIPRO, up to 30.2 µM. Its response was also confirmed selective for quinolones, when compared to its response against AMP. Further comparison to an immunosensor of similar design (adding antibodies instead of DNA gyrase) was made, revealing favourable features for the new biomimetic biosensor with 1.52 nM of limit of detection (LOD). Overall, the new approach presented herein is simple and effective for antibiotic detection, displaying a selective response against a given antibiotic group. The use of bacterial machinery as biorecognition element in biosensors may also provide a valuable tool to study the mechanism of action in bacterial cells of new drugs. This is especially important in the development of new drugs to fight bacterial resistance.
- In-situ production of Histamine-imprinted polymeric materials for electrochemical monitoring of fishPublication . Serrano, Verónica; Cardoso, Ana Rita; Diniz, Mário; Sales, Maria Goreti FerreiraHistamine (HIS) is a major public health problem due to its toxic properties. High levels can cause a chronic toxicity as poisoning and can be used as a signal of food hygiene. Thus, a new electrochemical sensor for HIS detection in fish is presented herein, prepared by tailoring a molecularly imprinted polymer (MIP) sensing material on a gold screen-printed electrode (Au-SPEs), in which the polymeric film was generated in-situ. This film was obtained by electropolymerizing aniline under conditions that preserved the chemical structure of HIS. Raman spectroscopy followed the chemical changes occurring at each stage of the electrode modification. The device performance was assessed by evaluating the changes in electron transfer properties of a standard redox probe [Fe(CN)6]4−/[Fe(CN)6]3−, by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). EIS was also used to calibrate the sensor, being the standard solutions prepared under different background media (electrolyte or a blank sample of fish extract). The device displayed a linear response from 500 nM to 1 mM, with a limit of detection of 210 nM, and a selective behaviour against tyramine, another amine related to fish degradation. The sensing system was further employed to monitor the HIS content in samples in different time points of storage at ambient temperature. The obtained results were in agreement with the ELISA method, while offering more reproducible data. In general, the optimized sensor allowed reproducible and accurate analysis of fish samples subject to degradation and was completely assembled in-situ, in a very simple and straightforward approach. The device is low cost and suitable for further adaptation to on-site analysis, as required in food control.
- An Innovative Approach for Tailoring Molecularly Imprinted Polymers for Biosensors—Application to Cancer Antigen 15-3Publication . Oliveira, Daniela dos Santos; Oliveira, Andreia Sofia Rodrigues; Mendonça, Patrícia Vitorino; Coelho, Jorge Fernando Jordão; Moreira, Felismina Teixeira Coelho; Sales, Maria Goreti Ferreira; Moreira, Felismina; DOS SANTOS OLIVEIRA, DANIELAThis work presents a novel approach for tailoring molecularly imprinted polymers (MIPs) with a preliminary stage of atom transfer radical polymerization (ATRP), for a more precise definition of the imprinted cavity. A well-defined copolymer of acrylamide and N,N′-methylenebisacrylamide (PAAm-co-PMBAm) was synthesized by ATRP and applied to gold electrodes with the template, followed by a crosslinking reaction. The template was removed from the polymer matrix by enzymatic/chemical action. The surface modifications were monitored via electrochemical impedance spectroscopy (EIS), having the MIP polymer as a non-conducting film designed with affinity sites for CA15-3. The resulting biosensor exhibited a linear response to CA15-3 log concentrations from 0.001 to 100 U/mL in PBS or in diluted fetal bovine serum (1000×) in PBS. Compared to the polyacrylamide (PAAm) MIP from conventional free-radical polymerization, the ATRP-based MIP extended the biosensor’s dynamic linear range 10-fold, improving low concentration detection, and enhanced the signal reproducibility across units. The biosensor demonstrated good sensitivity and selectivity. Overall, the work described confirmed that the process of radical polymerization to build an MIP material influences the detection capacity for the target substance and the reproducibility among different biosensor units. Extending this approach to other cancer biomarkers, the methodology presented could open doors to a new generation of MIP-based biosensors for point-of-care disease diagnosis.
- 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.