Search results for: Thiago Fabrizio Gavilano Farje

Authors: Shuo Li, Yannick Coffinier, Chann Lagadec, Fabrizio Cleri, Katsuhiko Nishiguchi, Akira Fujiwara, Soo Hyeon Kim, Nicolas Clément

Abstract:

The need for single cell detection and analysis techniques has increased in the past decades because of the heterogeneity of individual living cells, which increases the complexity of the pathogenesis of malignant tumors. In the search for early cancer detection, high-precision medicine and therapy, the technologies most used today for sensitive detection of target analytes and monitoring the variation of these species are mainly including two types. One is based on the identification of molecular differences at the single-cell level, such as flow cytometry, fluorescence-activated cell sorting, next generation proteomics, lipidomic studies, another is based on capturing or detecting single tumor cells from fresh or fixed primary tumors and metastatic tissues, and rare circulating tumors cells (CTCs) from blood or bone marrow, for example, dielectrophoresis technique, microfluidic based microposts chip, electrochemical (EC) approach. Compared to other methods, EC sensors have the merits of easy operation, high sensitivity, and portability. However, despite various demonstrations of low limits of detection (LOD), including aptamer sensors, arrayed EC sensors for detecting single-cell have not been demonstrated. In this work, a new technique based on 20-nm-thick nanopillars array to support cells and keep them at ideal recognition distance for redox-labeled aptamers grafted on the surface. The key advantages of this technology are not only to suppress the false positive signal arising from the pressure exerted by all (including non-target) cells pushing on the aptamers by downward force but also to stabilize the aptamer at the ideal hairpin configuration thanks to a confinement effect. With the first implementation of this technique, a LOD of 13 cells (with5.4 μL of cell suspension) was estimated. In further, the nanosupported cell technology using redox-labeled aptasensors has been pushed forward and fully integrated into a single-cell electrochemical aptasensor array. To reach this goal, the LOD has been reduced by more than one order of magnitude by suppressing parasitic capacitive electrochemical signals by minimizing the sensor area and localizing the cells. Statistical analysis at the single-cell level is demonstrated for the recognition of cancer cells. The future of this technology is discussed, and the potential for scaling over millions of electrodes, thus pushing further integration at sub-cellular level, is highlighted. Despite several demonstrations of electrochemical devices with LOD of 1 cell/mL, the implementation of single-cell bioelectrochemical sensor arrays has remained elusive due to their challenging implementation at a large scale. Here, the introduced nanopillar array technology combined with redox-labeled aptamers targeting epithelial cell adhesion molecule (EpCAM) is perfectly suited for such implementation. Combining nanopillar arrays with microwells determined for single cell trapping directly on the sensor surface, single target cells are successfully detected and analyzed. This first implementation of a single-cell electrochemical aptasensor array based on Brownian-fluctuating redox species opens new opportunities for large-scale implementation and statistical analysis of early cancer diagnosis and cancer therapy in clinical settings.

Keywords: bioelectrochemistry, aptasensors, single-cell, nanopillars

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Authors: Thiago R. Pereira

Abstract:

The philosophical hermeneutics is able to change the Brazilian Judiciary because of the understanding of the characteristics of the human being. It is impossible for humans, to be invested in the function of being a judge, making absolutely neutral decisions, but the philosophical hermeneutics can assist the judge making impartial decisions, based on the federal constitution. The normative legal positivism imagined a neutral judge, a judge able to try without any preconceived ideas, without allowing his/her background to influence him/her. When a judge arbitrates based on legal rules, the problem is smaller, but when there are no clear legal rules, and the judge must try based on principles, the risk of the decision is based on what they believe in. Solipsistically, this issue gains a huge dimension. Today, the Brazilian judiciary is independent, but there must be a greater knowledge of philosophy and the philosophy of law, partially because the bigger problem is the unpredictability of decisions made by the judiciary. Actually, when a lawsuit is filed, the result of this judgment is absolutely unpredictable. It is almost a gamble. There must be the slightest legal certainty and predictability of judicial decisions, so that people, with similar cases, may not receive opposite sentences. The relativism, since classical antiquity, believes in the possibility of multiple answers. Since the Greeks in in the sixth century before Christ, through the Germans in the eighteenth century, and even today, it has been established the constitution as the great law, the Groundnorm, and thus, the relativism of life can be greatly reduced when a hermeneut uses the Constitution as North interpretational, where all interpretation must act as the hermeneutic constitutional filter. For a current philosophy of law, that inside a legal system with a Federal Constitution, there is a single correct answer to a specific case. The challenge is how to find this right answer. The only answer to this question will be that we should use the constitutional principles. But in many cases, a collision between principles will take place, and to resolve this issue, the judge or the hermeneut will choose a solipsism way, using what they personally believe to be the right one. For obvious reasons, that conduct is not safe. Thus, a theory of decision is necessary to seek justice, and the hermeneutic philosophy and the linguistic turn will be necessary for one to find the right answer. In order to help this difficult mission, it will be necessary to use philosophical hermeneutics in order to find the right answer, which is the constitutionally most appropriate response. The constitutionally appropriate response will not always be the answer that individuals agree to, but we must put aside our preferences and defend the answer that the Constitution gives us. Therefore, the hermeneutics applied to Law, in search constitutionally appropriate response, should be the safest way to avoid judicial individual decisions. The aim of this paper is to present the science of law starting from the linguistic turn, the philosophical hermeneutics, moving away from legal positivism. The methodology used in this paper is qualitative, academic and theoretical, philosophical hermeneutics with the mission to conduct research proposing a new way of thinking about the science of law. The research sought to demonstrate the difficulty of the Brazilian courts to depart from the secular influence of legal positivism. Moreover, the research sought to demonstrate the need to think science of law within a contemporary perspective, where the linguistic turn, philosophical hermeneutics, will be the surest way to conduct the science of law in the present century.

Keywords: hermeneutic, right answer, solipsism, Brazilian judiciary

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Authors: Thiago E. Goto, Carla C. Lopes, Helena B. Nader, Anielle C. A. Silva, Noelio O. Dantas, José R. Siqueira Jr., Luciano Caseli

Abstract:

Quantum dots (QD) are semiconductor nanocrystals that can be employed in biological research as a tool for fluorescence imagings, having the potential to expand in vivo and in vitro analysis as cancerous cell biomarkers. Particularly, cadmium selenide (CdSe) magic-sized quantum dots (MSQDs) exhibit stable luminescence that is feasible for biological applications, especially for imaging of tumor cells. For these facts, it is interesting to know the mechanisms of action of how such QDs mark biological cells. For that, simplified models are a suitable strategy. Among these models, Langmuir films of lipids formed at the air-water interface seem to be adequate since they can mimic half a membrane. They are monomolecular films formed at liquid-gas interfaces that can spontaneously form when organic solutions of amphiphilic compounds are spread on the liquid-gas interface. After solvent evaporation, the monomolecular film is formed, and a variety of techniques, including tensiometric, spectroscopic and optic can be applied. When the monolayer is formed by membrane lipids at the air-water interface, a model for half a membrane can be inferred where the aqueous subphase serve as a model for external or internal compartment of the cell. These films can be transferred to solid supports forming the so-called Langmuir-Blodgett (LB) films, and an ampler variety of techniques can be additionally used to characterize the film, allowing for the formation of devices and sensors. With these ideas in mind, the objective of this work was to investigate the specific interactions of CdSe MSQDs with tumorigenic and non-tumorigenic cells using Langmuir monolayers and LB films of lipids and specific cell extracts as membrane models for diagnosis of cancerous cells. Surface pressure-area isotherms and polarization modulation reflection-absorption spectroscopy (PM-IRRAS) showed an intrinsic interaction between the quantum dots, inserted in the aqueous subphase, and Langmuir monolayers, constructed either of selected lipids or of non-tumorigenic and tumorigenic cells extracts. The quantum dots expanded the monolayers and changed the PM-IRRAS spectra for the lipid monolayers. The mixed films were then compressed to high surface pressures and transferred from the floating monolayer to solid supports by using the LB technique. Images of the films were then obtained with atomic force microscopy (AFM) and confocal microscopy, which provided information about the morphology of the films. Similarities and differences between films with different composition representing cell membranes, with or without CdSe MSQDs, was analyzed. The results indicated that the interaction of quantum dots with the bioinspired films is modulated by the lipid composition. The properties of the normal cell monolayer were not significantly altered, whereas for the tumorigenic cell monolayer models, the films presented significant alteration. The images therefore exhibited a stronger effect of CdSe MSQDs on the models representing cancerous cells. As important implication of these findings, one may envisage for new bioinspired surfaces based on molecular recognition for biomedical applications.

Keywords: biomembrane, langmuir monolayers, quantum dots, surfaces

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