Browsing by Author "Oliveira, Daniela"
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- Book of Abstracts of 5th Meeting of Medicinal Biotechnology (5MBtM) and 2nd Iberian Congress on Medicinal BiotechnologyPublication . Sieiro, Carmen; Pereira, Cassilda; Prudêncio, Cristina; Oliveira, Daniela; Teixeira, Dulce; Barros, Eduarda; Almeida, Joana; Cerqueira, Maria João; Louçano, Marta; Vieira, Mónica; Coelho, Pedro; Ferraz, Ricardo
- A cork based substrate coupled with artificial antibodies for point-of-care detection of pro-inflammatory cytokine biomarkersPublication . Correia, Bárbara; Oliveira, Daniela; Vulpe, Georgeta; Tavares, Ana P. M.; Sales, M. Goreti F.; Duarte, Abel J.; Sharma, Sanjiv; Moreira, Felismina T. C.Whilst single use point of care testing (PoCT) devices have transformed healthcare globally, there are major concerns over their environmental consequences. These concerns could be addressed by employing devices made of environmentally friendly materials. Herein, we report on the use of cork based PoCT devices. Cork is known to be fully biodegradable and can be easily recycled without producing toxic residues. We report on how a cork-based substrate coupled with a molecularly imprinted polymer (MIP) that serves as an “artificial antibody” can be used for point-of-care testing of the pro-inflammatory biomarker interleukin 6 (IL-6). The featured PoCT device has an electrochemical transducer that provides the desired clinical dynamic range for blood and can measure concentrations as low as 1 pg mL−1, indicating its usefulness in point of care measurements for monitoring pathological disorders, worldwide. In addition, it has a huge environmental impact as it can reduce the waste generated by other polymeric/ceramic carriers used for the same purpose.
- Development of an immunosensor based on quantum dots with different emissions for the detection of cancer biomarkersPublication . Oliveira, Daniela; Piloto, Ana Margarida L.(Introduction) Cancer is a global health challenge, where early detection is crucial to increasing survival rates [1]. Non-invasive, rapid, and affordable diagnostic methods are urgently needed. Immunosensors emerge as promising alternatives to traditional methods such as ELISA, allowing point-of-care (PoC) analyses [2]. This study introduces a fluorescent probe using cadmium telluride quantum dots (CdTe@MPA QDs) to detect the pancreatic cancer biomarker CA 19-9. With the increasing CA 19-9 concentrations, the fluorescence intensity of green-, orange-, and red-emitting QDs@conjugates was reduced, creating a visible colour gradient under a 365 nm UV lamp. The method, suitable for serum analysis, is highly sensitive and selective, with a detection range of 0.31 to 501.9 U mL-1 and analyzed using ImageJ software. These immunosensors offer significant potential for cancer biomarker detection in clinical diagnostics.
- Electrochemical miRNA-34a-based biosensor for the diagnosis of Alzheimer’s diseasePublication . Pereira, Raquel L.; Oliveira, Daniela; Pêgo, Ana P.; Santos, Sofia D.; Moreira, Felismina T.C.Alzheimer's disease (AD) is the most common dementia type and a leading cause of death and disability in the elderly. Diagnosis is expensive and invasive, urging the development of new, affordable, and less invasive diagnostic tools. The identification of changes in the expression of non-coding RNAs prompts the development of diagnostic tools to detect disease-specific blood biomarkers. Building on this idea, this work reports a novel electrochemical microRNA (miRNA) biosensor for the diagnosis of AD, based on carbon screen-printed electrodes (C-SPEs) modified with two gold nanostructures and a complementary anti-miR-34a oligonucleotide probe. This biosensor showed good target affinity, reflected on a 100 pM to 1 μM linearity range and a limit of detection (LOD) of 39 pM in buffer and 94 aM in serum. Moreover, the biosensor’s response was not affected by serum compounds, indicating selectivity for miR-34a. The biosensor also detected miR-34a in the cell culture medium of a common AD model, stimulated with a neurotoxin to increase miR-34a secretion. Overall, the proposed biosensor makes a solid case for the introduction of a novel, inexpensive, and minimally invasive tool for the early diagnosis of AD, based on the detection of a circulating miRNA overexpressed in this pathology.
- Electrochemical miRNA-34a-based biosensor for the diagnosis of Alzheimer’s diseasePublication . Pereira, Raquel L.; Oliveira, Daniela; Pêgo, Ana P.; Santos, Sofia D.; Moreira, Felismina T.C.Alzheimer's disease (AD) is the most common dementia type and a leading cause of death and disability in the elderly. Diagnosis is expensive and invasive, urging the development of new, affordable, and less invasive diagnostic tools. The identification of changes in the expression of non-coding RNAs prompts the development of diagnostic tools to detect disease-specific blood biomarkers. Building on this idea, this work reports a novel electrochemical microRNA (miRNA) biosensor for the diagnosis of AD, based on carbon screen-printed electrodes (C-SPEs) modified with two gold nanostructures and a complementary anti-miR-34a oligonucleotide probe. This biosensor showed good target affinity, reflected on a 100 pM to 1 μM linearity range and a limit of detection (LOD) of 39 pM in buffer and 94 aM in serum. Moreover, the biosensor’s response was not affected by serum compounds, indicating selectivity for miR-34a. The biosensor also detected miR-34a in the cell culture medium of a common AD model, stimulated with a neurotoxin to increase miR-34a secretion. Overall, the proposed biosensor makes a solid case for the introduction of a novel, inexpensive, and minimally invasive tool for the early diagnosis of AD, based on the detection of a circulating miRNA overexpressed in this pathology.
- An electrochemically synthesized molecularly imprinted polymer for highly selective detection of breast cancer biomarker CA 15-3: a promising point-of-care biosensorPublication . Oliveira, Daniela; Barcelay, Yonny Romaguera; Moreira, Felismina T. C.In this study, a molecularly imprinted polymer film (MIP) was prepared on the surface of a disposable carbon screen-printed electrode (C-SPE) using (3-acrylamidopropyl)trimethylammonium chloride (AMPTMA) as a functional monomer and the cancer biomarker carbohydrate antigen 15-3 (CA 15-3) as a template. The MIP was synthesized by in situ electropolymerization (ELP) of the AMPTMA monomer in the presence of the CA 15-3 protein on the C-SPE surface. The target was subsequently removed from the polymer matrix by the action of proteinase K, resulting in imprinted cavities with a high affinity for CA 15-3. Electrochemical techniques such as cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to characterize the different phases of the sensor assembly. Chemical and morphological analysis was performed using RAMAN and scanning electron microscopy (SEM). CA 15-3 was successfully detected in a wide working range from 0.001 U mL−1 to 100 U mL−1 with a correlation coefficient (R2) of 0.994 in 20 min. The MIP sensor showed minimal interference with other cancer proteins (CEA and CA 125). Overall, the developed device provides a rapid, sensitive, and cost-effective response in the detection of CA 15-3. Importantly, this comprehensive approach appears suitable for point-of-care (PoC) use, particularly in a clinical context.
- An electrochemically synthesized molecularly imprinted polymer for highly selective detection of breast cancer biomarker CA 15-3: a promising point-of-care biosensorPublication . Oliveira, Daniela; Romaguera Barcelay, Yonny; Moreira, FelisminaIn this study, a molecularly imprinted polymer film (MIP) was prepared on the surface of a disposable carbon screen-printed electrode (C-SPE) using (3-acrylamidopropyl)trimethylammonium chloride (AMPTMA) as a functional monomer and the cancer biomarker carbohydrate antigen 15-3 (CA 15-3) as a template. The MIP was synthesized by in situ electropolymerization (ELP) of the AMPTMA monomer in the presence of the CA 15-3 protein on the C-SPE surface. The target was subsequently removed from the polymer matrix by the action of proteinase K, resulting in imprinted cavities with a high affinity for CA 15-3. Electrochemical techniques such as cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to characterize the different phases of the sensor assembly. Chemical and morphological analysis was performed using RAMAN and scanning electron microscopy (SEM). CA 15-3 was successfully detected in a wide working range from 0.001 U mL−1 to 100 U mL−1 with a correlation coefficient (R2) of 0.994 in 20 min. The MIP sensor showed minimal interference with other cancer proteins (CEA and CA 125). Overall, the developed device provides a rapid, sensitive, and cost-effective response in the detection of CA 15-3. Importantly, this comprehensive approach appears suitable for point-of-care (PoC) use, particularly in a clinical context.
- Enhanced detection with prussian blue-based nanocubes: a novel electrochemical biosensor for bovine serum albumin analysis: microfluidic integrationPublication . Santos, Andréa dos; Oliveira, Daniela; Martins, Gabriela; Moreira, Felismina T.C.(Introduction) Early diagnosis of biomarkers is crucial for optimizing treatments and increasing survival rates. Direct and cost-effective assays are important for point-of-care (PoC) testing. Bovine serum albumin (BSA) monitoring indicates liver and kidney function and aids in the assessment of diseases such as cirrhosis and chronic kidney disease. An innovative electrochemical biosensor was developed in which a platinum electrode was decorated with manganese-based Prussian blue nanocubes (PB-NC). A molecularly imprinted polymer (MIP) was created on the electrode surface by electropolymerization of phenol in the presence of BSA using cyclic voltammetry (CV). The template was removed with proteinase K and a mixture of methanol and acetic acid. Square wave voltammetry (SWV) characterized the performance of the sensor, which exhibited high sensitivity and allowed detection of BSA at nanomolar levels.
- FASL polymorphism is associated with response to bacillus Calmette-Guérin immunotherapy in bladder cancerPublication . Lima, Luís; Ferreira, José Alexandre; Tavares, Ana; Oliveira, Daniela; Morais, António; Videira, Paula; Medeiros, Rui; Santos, LúcioObjective Deregulation of FAS/FASL system may lead to immune escape and influence bacillus Calmette-Guérin (BCG) immunotherapy outcome, which is currently the gold standard adjuvant treatment for high-risk non–muscle invasive bladder tumors. Among other events, functional promoter polymorphisms of FAS and FASL genes may alter their transcriptional activity. Therefore, we aim to evaluate the role of FAS and FASL polymorphisms in the context of BCG therapy, envisaging the validation of these biomarkers to predict response. Patients and methods DNA extracted from peripheral blood from 125 patients with bladder cancer treated with BCG therapy was analyzed by Polymerase Chain Reaction—Restriction Fragment Length Polymorphism for FAS-670 A/G and FASL-844 T/C polymorphisms. FASL mRNA expression was analyzed by real-time Polymerase Chain Reaction. Results Carriers of FASL-844 CC genotype present a decreased recurrence-free survival after BCG treatment when compared with FASL-844 T allele carriers (mean 71.5 vs. 97.8 months, P = 0.030) and have an increased risk of BCG treatment failure (Hazard Ratio = 1.922; 95% Confidence Interval: [1.064–3.471]; P = 0.030). Multivariate analysis shows that FASL-844 T/C and therapeutics scheme are independent predictive markers of recurrence after treatment. The evaluation of FASL gene mRNA levels demonstrated that patients carrying FASL-844 CC genotype had higher FASL expression in bladder tumors (P = 0.0027). Higher FASL levels were also associated with an increased risk of recurrence after BCG treatment (Hazard Ratio = 2.833; 95% Confidence Interval: [1.012–7.929]; P = 0.047). FAS-670 A/G polymorphism analysis did not reveal any association with BCG therapy outcome. Conclusions Our results suggest that analysis of FASL-844 T/C, but not FAS-670 A/G polymorphisms, may be used as a predictive marker of response to BCG immunotherapy.
- Macrophages as biomarkers in BCG treatment response in bladder cancerPublication . Oliveira, Daniela; Ferreira, José; Prudêncio, Cristina; Lima, LuísBladder cancer is a common urologic cancer and the majority has origin in the urothelium. Patients with intermediate and high risk of recurrence/progression bladder cancer are treated with intravesical instillation with Bacillus Calmette-Guérin, however, approximately 30% of patients do not respond to treatment. At the moment, there are no accepted biomarkers do predict treatment outcome and an early identification of patients better served by alternative therapeutics. The treatment initiates a cascade of cytokines responsible by recruiting macrophages to the tumor site that have been shown to influence treatment outcome. Effective BCG therapy needs precise activation of the Th1 immune pathway associated with M1 polarized macrophages. However, tumor-associated macrophages (TAMs) often assume an immunoregulatory M2 phenotype, either immunosuppressive or angiogenic, that interfere in different ways with the BCG induced antitumor immune response. The M2 macrophage is influenced by different microenvironments in the stroma and the tumor. In particular, the degree of hypoxia in the tumors is responsible by the recruitment and differentiation of macrophages into the M2 angiogenic phenotype, suggested to be associated with the response to treatment. Nevertheless, neither the macrophage phenotypes present nor the influence of localization and hypoxia have been addressed in previous studies. Therefore, this work devoted to study the influence of TAMs, in particular of the M2 phenotype taking into account their localization (stroma or tumor) and the degree of hypoxia in the tumor (low or high) in BCG treatment outcome. The study included 99 bladder cancer patients treated with BCG. Tumors resected prior to treatment were evaluated using immunohistochemistry for CD68 and CD163 antigens, which identify a lineage macrophage marker and a M2-polarized specific cell surface receptor, respectively. Tumor hypoxia was evaluated based on HIF-1α expression. As a main finding it was observed that a high predominance of CD163+ macrophage counts in the stroma of tumors under low hypoxia was associated with BCG immunotherapy failure, possibly due to its immunosuppressive phenotype. This study further reinforces the importance the tumor microenvironment in the modulation of BCG responses.
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