Plasma Activation of Microplates Optimized for One-Step Reagent-Free Immobilization of DNA and Protein

Activated microplates are widely used in biological assays and cell culture to immobilize biomolecules, either through passive physical adsorption or covalent cross-linking. Covalent attachment gives greater stability in complex biological mixtures. However, current multistep chemical activation methods add complexity and cost, require specific functional groups, and can introduce cytotoxic chemicals that affect downstream cellular applications. Here, we show a method for one-step linker-free activation of microplates by energetic ions from plasma for covalent immobilization of DNA and protein. Two types of energetic ion plasma treatment were shown to be effective: plasma immersion ion implantation (PIII) and plasma-activated coating (PAC). This is the first time that PIII and PAC have been reported in microwell plates with nonflat geometry. We confirm that the plasma treatment generates radical-activated surfaces at the bottom of wells despite potential shadowing from the walls. Comprehensive surface characterization studies were used to compare the PIII and PAC microplate surface composition, wettability, radical density, optical properties, stability, and biomolecule immobilization density. PAC plates were found to have more nitrogen and lower radical density and were more hydrophobic and more stable over 3 months than PIII plates. Optimal conditions were obtained for high-density DNA (PAC, 0 or 21% nitrogen, pH 3–4) and streptavidin (PAC, 21% nitrogen, pH 5–7) binding while retaining optical properties required for typical high-throughput biochemical microplate assays, such as low autofluorescence and high transparency. DNA hybridization and protein activity of immobilized molecules were confirmed. We show that PAC activation allows for high-density covalent immobilization of functional DNA and protein in a single step on both 96- and 384-well plates without specific linker chemistry. These microplates could be used in the future to bind other user-selected ligands in a wide range of applications, for example, for solid phase polymerase chain reaction and stem cell culture and differentiation

L'essai d'infiltration couplé à un traçage non réactif : un outil pour évaluer le transfert des polluants dans la zone non saturée des sols

International audienceThe unsaturated zone of the soil is generally of great importance in order to describe the fate of contaminants in the subsurface. Because of its central position exchanging with the other compartments of the environment (atmosphere, biosphere and groundwater), the unsaturated zone is the place of highly transient water flow conditions. At the scale of a spill at the soil surface at an industrial unit, the heterogeneous distribution of water flow and contaminants transport properties in the unsaturated zone often results in a wide range of impact times and concentrations to groundwater. A research project named TRANSAT has then been conducted by the group composed of CEA / BURGEAP / LTHE / EDF with the funding of the National Research Agency. The edited guidelines documentation gives technical and methodological recommendations aiming to specify when and how the unsaturated zone needs to be accounted for and what type of characterization and modelling tools match the spill scenario. Among the available methods, a special interest has grown towards the coupled large diameter ring infiltration and conservative tracer test, proven to be the only tool capable of measuring the impact time and concentration in the groundwater at the scale of the site spill area and accounting for unsaturated zone characteristics: heterogeneity and capillary retention / dispersion effects.La zone non saturée du sol (ZNS) joue généralement un rôle important dans le devenir des polluants dans le milieu souterrain. Étant située en position centrale dans les échanges avec les autres compartiments de l’environnement (atmosphère, biosphère et eau souterraine), la ZNS est soumise à des conditions d’écoulement d’eau fortement transitoires. À l’échelle d’un déversement en surface au droit d’une unité industrielle, l’hétérogénéité des propriétés d’écoulement d’eau et de transport des polluants dans la ZNS se traduit généralement par une large disparité des temps d’arrivée et des concentrations des polluants au toit de la nappe. Dans ce contexte, le consortium CEA / BURGEAP / LTHE / EDF a mené un projet de recherche & développement co-financé par l’Agence Nationale de la Recherche : TRANSAT. Le guide issu de ce projet décrit des recommandations techniques et méthodologiques visant à préciser quand et comment prendre en compte la ZNS et avec quels outils de caractérisation et de modélisation en fonction du scenario de déversement. Parmi les techniques disponibles, une attention particulière a été portée sur l’essai d’infiltration gros diamètre couplé à un traçage en ZNS, seule méthode permettant de mesurer, à l’échelle de la zone de déversement du site, les temps et concentrations d’impact à la nappe en intégrant les hétérogénéités et les effets de rétention capillaire/dispersion dans la ZNS

Características associadas à virulência de Enterococcus faecalis isolados de casos clínicos

Thirty-two clinical isolates of Enterococcus faecalis were screened for virulence factors. Twenty-four (75%) isolates produced hemolysin on Mueller-Hinton blood agar plates with sheep erythrocytes. However, the cell free heat-stable hemolysin was detected in all isolates (100%) of E. faecalis when grown in BHI-GA (BHI medium supplemented with 1% glucose and 0.03% L-arginine), but not in BHI broth alone. Twenty-four isolates (75%) produced caseinase and 23 (71.9%) lipase, but none of the isolates produced gelatinase. Fifteen (46.9%) culture filtrates caused rounding and membrane alterations with blebbing formation followed by death in HeLa and HEp-2 cells, but not in Vero cells. Thirteen isolates (40.6%) agglutinated rabbit erythrocytes, but did not produce hemagglutination in other bloods, containing or not 1% D-manose. Sixteen (50%) E. faecalis isolates adhered to HeLa cells and thirteen (40.6%) to HEp-2 cells, but all isolates adhered to polypropylene microtiter plates, indicating that clinical E. faecalis possess the ability to form biofilm in vitro. All the isolates were resistant to the bactericidal action of normal serum and did not produce aerobactin. These findings suggest that adherence and consequently biofilm formation on ephitelial host cells are the first steps in the E. faecalis virulence and that hemolysin, lipase, caseinase and other virulence factors act as causative of human epithelial cell damages.Foram estudados os fatores de virulência de trinta e duas amostras de Enterococcus faecalis, isolados de casos clínicos. Vinte e quatro amostras (75%) produziram hemolisina em ágar sangue preparado com hemácias de carneiro. No sobrenadante da cultura em BHI nenhuma amostra produziu hemolisina, no entanto quando cultivadas em meio BHI suplementado com 1% de glucose e 0,03% de L-arginina (BHI-GA), 100% das amostras lisaram hemácias de carneiro. Vinte e quatro (75%) amostras produziram caseinase e 23 (71,9%) lipase, mas nenhuma amostra produziu gelatinase. Dezesseis (46,9%) causaram arredondamento e alteração na membrana das células, com formação de vesículas e, em seguida, a morte das células HeLa e HEp-2. Treze amostras (40,6) aglutinaram eritrócitos de coelhos, mas não aglutinaram outros eritrócitos na presença ou na ausência de 1% de D-manose. Dezesseis (50%) aderiram em células HeLa e 13 (40,6%) em células HEp-2, mas todas as amostras de E. faecalis aderiram na microplaca de polipropileno, indicando que E. faecalis isolados de casos clínicos possuem capacidade de formar biofilme in vitro. Todos os isolados mostraram-se resistentes à ação bactericida do soro normal e não produziram aerobactina. Esses resultados sugerem que, inicialmente, a colonização ou infecção por E. faecalis ocorre pela aderência e formação de biofilme nas células epiteliais e a produção de hemolisina, lípase e caseinase pode atuar como fatores de virulência na infecção por E. faecalis.230236Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES

Adult breast, lung, pancreatic, upper and lower gastrointestinal cancer patients with hospitalized venous thromboembolism in the national French hospital discharge database

Abstract Background Venous thromboembolism (VTE) and cancer are strongly associated. In France, evidence on patients with pancreatic, upper GI [gastrointestinal], lower GI, lung, or breast cancer-associated VTE and their hospital management is limited. The aims of this study were to provide data on the number of hospitalized VTE events among cancer patients, the patients’ characteristics, and their hospital management to estimate the burden of disease and the hospital burden of cancer-related VTE and to provide guidance on research. Methods This longitudinal, observational, and retrospective study was based on the comprehensive hospital discharge database (PMSI). Adult patients (≥ 18 years old) hospitalized with a cancer of interest in 2016 and hospitalized (within 2 years with VTE (captured a as a principal, related, or significant associated diagnosis) were included in the study. Results We identified 340,946 cancer patients, of which 7.2% (24,433 patients) were hospitalized with VTE. The proportions of hospitalized VTE were 14.6% (3,237) for patients with pancreatic cancer, 11.2% (8,339) for lung cancer, 9.9% (2,232) for upper GI cancer, 6.7% (7,011) for lower GI cancer, and 3.1% (3,614) for breast cancer. Around two thirds of cancer patients with a hospitalized VTE had active cancer (with metastases and/or receiving chemotherapy during the six months prior to the index date): from 62% of patients with pancreatic cancer to 72% with breast cancer. Around a third of patients were admitted to the hospital through the emergency room, up to 3% of patients stayed in an intensive care unit. The average length of stay ranged from 10 (breast cancer) to 15 days (upper GI cancer). Nine (lower GI cancer) to 18% (pancreatic cancer) of patients died during the VTE hospital stay. Conclusions The burden of cancer-associated VTE is substantial, both in terms of the number of patients affected and in the hospital use. These findings offer guidance on future research on VTE prophylaxis in a very high-risk population, particularly in patients with active cancer

Identification of LiNbO3, LiNb3O8 and Li3NbO4 phases in thin films synthesized with different deposition techniques by means of XRD and Raman spectroscopy

Équipe 401 : Nanomatériaux pour la vie et développement durableInternational audiencePhase composition of epitaxial/textured LiNbO3 films on sapphire substrates, grown by pulsed laser deposition, atmospheric pressure metal organic chemical vapor deposition and pulsed injection metal organic chemical vapor deposition was studied by conventional x-ray diffraction techniques. Raman spectroscopy, being highly sensitive to the symmetry of materials, was used as a countercheck in the compositional analysis. The wavenumbers of Raman modes of LiNb3O8 and Li3NbO4 phases were identified from Raman spectra of synthesized powders. Asymmetry of profiles of x-ray diffraction reflections of LiNbO3 films was studied. This asymmetry may have different origins which consequently may result in misleading conclusions about phase composition of textured LiNbO3 films

Residual stresses and clamped thermal expansion in LiNbO3 and LiTaO3 thin films

International audienc