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Nat Commun ; 11(1): 2668, 2020 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-32472057


Electrochemiluminescence (ECL) is a powerful transduction technique with a leading role in the biosensing field due to its high sensitivity and low background signal. Although the intrinsic analytical strength of ECL depends critically on the overall efficiency of the mechanisms of its generation, studies aimed at enhancing the ECL signal have mostly focused on the investigation of materials, either luminophores or coreactants, while fundamental mechanistic studies are relatively scarce. Here, we discover an unexpected but highly efficient mechanistic path for ECL generation close to the electrode surface (signal enhancement, 128%) using an innovative combination of ECL imaging techniques and electrochemical mapping of radical generation. Our findings, which are also supported by quantum chemical calculations and spin trapping methods, led to the identification of a family of alternative branched amine coreactants, which raises the analytical strength of ECL well beyond that of present state-of-the-art immunoassays, thus creating potential ECL applications in ultrasensitive bioanalysis.

Anal Bioanal Chem ; 411(19): 4375-4382, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31020369


The use of electrochemiluminescence (ECL), i.e., chemiluminescence triggered by electrochemical stimulus, as emitting light source for microscopy is an emerging approach with different applications ranging from the visualization of nanomaterials to cell mapping. In this trend article, we give an overview of the state of the art in this new field with the purpose to illustrate all the possible applications so far explored as well as describing the mechanism underlying this transduction technique. The results discussed here would highlight the great potential of the combination between ECL and microscopy and how this marriage can turn into an innovative approach with specific application in analytical sciences. Graphical abstract.

Técnicas Eletroquímicas/métodos , Luminescência , Medições Luminescentes/métodos , Microscopia/métodos , Eletrodos , Nanoestruturas , Análise de Célula Única
Anal Bioanal Chem ; 408(25): 7085-94, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27039202


Nanoelectrode arrays (NEAs) are increasingly applied for a variety of electroanalytical applications; however, very few studies dealt with the use of NEAs as an electrochemical generator of electrogenerated chemiluminescence (ECL). In the present study, arrays of nanodisc and nanoband electrodes with different dimensions and inter-electrode distances were fabricated by e-beam lithography on a polycarbonate layer deposited on boron-doped diamond (BDD) substrates. In particular, NEAs with 16 different geometries were fabricated on the same BDD sample substrate obtaining a multiple nanoelectrode and ultramicroelectrode array platform (MNEAP). After electrochemical and morphological characterization, the MNEAP was used to capture simultaneously with a single image the characteristic behaviour of ECL emission from all the 16 arrays. Experiments were performed using Ru(bpy)3 (2+) as the ECL luminophore and tri-n-propylamine (TPrA) as the co-reactant. With a relatively limited number of experiments, such an imaging procedure allowed to study the role that geometrical and mechanistic parameters play on ECL generation at NEAs. In particular, at high concentrations of TPrA, well-separated individual ECL spots or bands revealed an ECL signal which forms a pattern matching the nanofabricated structure. The analysis of the imaging data indicated that the thickness of the ECL-emitting zone at each nanoelectrode scales inversely with the co-reactant concentration, while significantly stronger ECL signals were detected for NEAs operating under overlap conditions.

Nat Genet ; 47(6): 607-14, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25938942


Glucocorticoids are universally used in the treatment of acute lymphoblastic leukemia (ALL), and resistance to glucocorticoids in leukemia cells confers poor prognosis. To elucidate mechanisms of glucocorticoid resistance, we determined the prednisolone sensitivity of primary leukemia cells from 444 patients newly diagnosed with ALL and found significantly higher expression of CASP1 (encoding caspase 1) and its activator NLRP3 in glucocorticoid-resistant leukemia cells, resulting from significantly lower somatic methylation of the CASP1 and NLRP3 promoters. Overexpression of CASP1 resulted in cleavage of the glucocorticoid receptor, diminished the glucocorticoid-induced transcriptional response and increased glucocorticoid resistance. Knockdown or inhibition of CASP1 significantly increased glucocorticoid receptor levels and mitigated glucocorticoid resistance in CASP1-overexpressing ALL. Our findings establish a new mechanism by which the NLRP3-CASP1 inflammasome modulates cellular levels of the glucocorticoid receptor and diminishes cell sensitivity to glucocorticoids. The broad impact on the glucocorticoid transcriptional response suggests that this mechanism could also modify glucocorticoid effects in other diseases.

Proteínas de Transporte/metabolismo , Caspase 1/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/enzimologia , Receptores de Glucocorticoides/metabolismo , Adolescente , Antineoplásicos Hormonais/farmacologia , Sequência de Bases , Criança , Pré-Escolar , Metilação de DNA , Resistencia a Medicamentos Antineoplásicos , Ensaios de Seleção de Medicamentos Antitumorais , Regulação Leucêmica da Expressão Gênica , Células HEK293 , Humanos , Lactente , Recém-Nascido , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR , Recidiva Local de Neoplasia/enzimologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Prednisolona/farmacologia , Proteólise , Transcrição Genética , Células Tumorais Cultivadas , Regulação para Cima