Particle emission rates during electrostatic spray deposition of TiO2 nanoparticle-based photoactive coating

A new method for the covalent and specific labeling of fusion proteins of carrier proteins (CPs) with small organic molecules has been developed in this work. This technology combines the convenience of expressing genetically tagged reporter proteins with the versatility of synthetic organic molecules. Moreover it promises to overcome some of the limitations of the currently used approaches. The method is based on the posttranslational modification of CPs by phosphopantetheine transferase (PPTase). In this reaction, the 4'-phosphopantetheine group of coenzyme A (CoA) is transferred to a serine residue of CP by PPTase. The PPTase can also use as substrates CoA derivatives that are modified in the thiol moiety by fluorophores or affinity reporter groups that are transferred to CP by PPTase in a covalent and irreversible manner. In this work, several CoA derivatives were synthesized by coupling of CoA with reporter groups functionalized by maleimide. The labeling method using the acyl carrier protein (ACP) and the PPTase (AcpS) from E. coli was applied to the in vitro labeling of purified proteins or in E. coli and yeast lysates, but also to the labeling of proteins expressed on cell surfaces of yeast and mammalian cells. The labeling reaction is fast, specific and quantitative. Pulse-chase labeling experiments with different fluorophores allowed the visualization of different protein generations on yeast cell surfaces. Thus, the method was demonstrated to be attractive for fluorescence microscopy. The second objective was to create a system for the selective labeling of different CPs with different CoA derivatives in the same sample, which requires PPTases with different specificities. The labeling must be performed sequentially, in order that each CP is labeled with only one CoA derivative. The pair peptidyl carrier protein (PCP) from B. brevis and the PPTase from B. subtilis (Sfp) was chosen as counterpart of the pair ACP / AcpS from E. coli. AcpS that is specific towards ACP is used for the first labeling reaction, and after a washing step to remove excess of substrate, the second labeling is performed with Sfp which is promiscuous. The system was successfully tested in vitro in solution and with proteins immobilized on microarrays, and on the surface of yeast and mammalian cells. Finally, the last objective was to reduce the size of the carrier protein (∼ 80 amino acids) to a minimal motif that is efficiently recognized by the PPTase. ACP and PCP were truncated before and after helix II whose residues are involved in the recognition by AcpS and Sfp. The fragments of ACP (aa 27-50) and PCP (aa 37-59) were labeled by AcpS and Sfp respectively, but the kinetics of labeling was slow. Two libraries were created with randomization of the six amino acids around the modified serine. Selections were performed using a phage display system based on the phagemid technology. Mt1 (32 aa) was modified by AcpS at the same rate as wild type ACP. Additional truncations of mt1 sequence yielded mt1.4 (16 aa) that was efficiently recognized by AcpS and weakly by Sfp. In conclusion, this labeling method should become an important tool for studies of cell surface proteins as well as for in vitro applications

Comparison of geometrical layouts for a multi-box aerosol model from a single-chamber dispersion study

Models are increasingly used to estimate and pre-emptively calculate the occupational exposure of airborne released particulate matter. Typical two-box models assume instant and fully mixed air volumes, which can potentially cause issues in cases with fast processes, slow air mixing, and/or large volumes. In this study, we present an aerosol dispersion model and validate it by comparing the modelled concentrations with concentrations measured during chamber experiments. We investigated whether a better estimation of concentrations was possible by using different geometrical layouts rather than a typical two-box layout. A one-box, two-box, and two three-box layouts were used. The one box model was found to underestimate the concentrations close to the source, while overestimating the concentrations in the far field. The two-box model layout performed well based on comparisons from the chamber study in systems with a steady source concentration for both slow and fast mixing. The three-box layout was found to better estimate the concentrations and the timing of the peaks for fluctuating concentrations than the one-box or two-box layouts under relatively slow mixing conditions. This finding suggests that industry-relevant scaled volumes should be tested in practice to gain more knowledge about when to use the two-box or the three-box layout schemes for multi-box models

Nanotitanium dioxide toxicity in mouse lung is reduced in sanding dust from paint

<p>Abstract</p> <p>Background</p> <p>Little is known of how the toxicity of nanoparticles is affected by the incorporation in complex matrices. We compared the toxic effects of the titanium dioxide nanoparticle UV-Titan L181 (NanoTiO₂), pure or embedded in a paint matrix. We also compared the effects of the same paint with and without NanoTiO₂.</p> <p>Methods</p> <p>Mice received a single intratracheal instillation of 18, 54 and 162 μg of NanoTiO₂or 54, 162 and 486 μg of the sanding dust from paint with and without NanoTiO₂. DNA damage in broncheoalveolar lavage cells and liver, lung inflammation and liver histology were evaluated 1, 3 and 28 days after intratracheal instillation. Printex 90 was included as positive control.</p> <p>Results</p> <p>There was no additive effect of adding NanoTiO₂to paints: Therefore the toxicity of NanoTiO₂was reduced by inclusion into a paint matrix. NanoTiO₂induced inflammation in mice with severity similar to Printex 90. The inflammatory response of NanoTiO₂and Printex 90 correlated with the instilled surface area. None of the materials, except of Printex 90, induced DNA damage in lung lining fluid cells. The highest dose of NanoTiO₂caused DNA damage in hepatic tissue 1 day after intratracheal instillation. Exposure of mice to the dust from paints with and without TiO₂was not associated with hepatic histopathological changes. Exposure to NanoTiO₂or to Printex 90 caused slight histopathological changes in the liver in some of the mice at different time points.</p> <p>Conclusions</p> <p>Pulmonary inflammation and DNA damage and hepatic histopathology were not changed in mice instilled with sanding dust from NanoTiO₂paint compared to paint without NanoTiO₂. However, pure NanoTiO₂caused greater inflammation than NanoTiO₂embedded in the paint matrix.</p