5 research outputs found
New Pd/Pt on Mg/Al basic mixed oxides for vapor phase hydrogenation and hydrogenolysis of naphthalene
No full textNew Pd/Pt catalysts supported on a basicMg/Al mixed oxide obtained by calcination of a commercial hydrotalcite (SASOL, D) have been fully characterized by XRD, SEM, and FTIR and investigated in the vapor-phase hydrogenation of naphthalene in order to put in evidence
the role of the Pd/Pt active phase and the acidity of the support on the hydrogenolysis/ring-opening reaction as well as the thio-tolerance of the catalysts. After calcination the hydrotalcite support (HT) gave rise to a Mg/Al mixed oxide with high surface area and a “brain-like” surface morphology. The IR spectra after adsorption of CO over the reduced samples showed the interaction between bimetallic particles and Mg–Al(O) basic support. The main band in the range 2080–2070 cm−1 is due to on-top CO adsorbed over Pd0 or Pt0, while the complex absorption below 2000 cm−1 is due to CO species bridging over Pd0. All the samples showed mainly a hydrogenation activity, highlighting the role of the support acidity in the ring-opening reactions to high molecular weight (HMW) products, having a boosting effect on the cetane number of the fraction obtained. A significant increase in the amount of HMWproducts was obtained by decreasing the Pd/Pt ratio, showing also the role of hydrogenolysis reactions attributable mainly to Pt, thus suggesting a hydrogenolysis/ring-opening mechanism. Furthermore, the Pd/Pt on basic oxide catalysts did not give rise to useless low molecular weight (LMW) cracking compounds, which thus formed only on
highly acid sites. Finally, these catalysts, investigated by feeding increasing amounts of dibenzothiophene (DBT), showed an almost constant activity up to 3000 wt ppm of DBT. This surprising result pointed out the intrinsic thio-resistance of the Pd/Pt pair, regardless of a possible
contribution of the acid sites of the support, and is mainly attributed to its high hydrodesulfurization (HDS) activity
The life marker chip : potential use of aptamers against small molecules and consideration of instrument planetary protection
No full textThe Life Marker Chip (LMC) instrument was developed with the aim to detect evidence of life on Mars. The detection was based on an inhibition immunoassay. In this work aptamers were evaluated as potential alternative to antibodies for the LMC. Aptamers were synthetic oligonucleotides able to bind specifically with high affinity to a wide range of target molecules, and have been also integrated as bioreceptors in several detection instruments. The generation of new aptamers against two small molecules using the FluMag-SELEX method was tested and was verified the adaptability of pre-existing aptamers against small targets to the LMC assay type. Based on the fact that the LMC was going to be integrated into the space programme ExoMars, it was also implemented into a small scale experiment the Planetary Protection and Contamination Control requirements found on a life-search mission. In addition to that aptamers compatibility with a sterilisation procedure used in life-search missions was also tested. Furthermore because of the nature of the small molecules studied, multiple analytical chemistry techniques were assessed to verify covalent chemistry surface immobilisation.
Within the project timeline it was not possible to achieve a full aptamer generation process but it was possible to understand the methodology behind the procedure and give input for future work. It was found that the direct implementation of existing aptamers against small molecules into the LMC assay was not successful. It was also seen that in the case of aptamer integration onto the LMC some assay changes would probably have to be made. This information was very useful to understand if aptamers could be an alternative to antibodies and be implemented directly into the LMC. It was found that aptamers survived the preliminary sterilisation method applied, which might open the possibility of making aptamers convenient space bioreceptors, reducing time and costs of instrument Planetary Protection implementation. In conclusion aptamers were not straightforward alternatives to antibodies for the LMC because aptamers interacted differently with their targets in comparison to antibodies, particularly with small molecules. Also the biochemical simplicity of the small molecule targets introduced difficulties in aptamers generation that more complex targets would have not. Although aptamers shown incompatibility with the LMC assay format against small targets, they presented resilience to a sterilisation procedure implemented on space missions which could lead to the development of more robust bioreceptors for space missions. This information was helpful in understanding which assay formats were better for detection of small molecules using aptamers and that might contribute for future assay choices applied in detection instruments. It was also possible to make recommendations for the LMC regarding design and validation methods used in life-search missions based on the lessons learn from the developed of a small scale experiment.
The developed work was presented at conferences and mentioned in an article journal, and in that way contributed to the knowledge of the space community in general
Standardized development of microarray technology via substrate-independent surface coatings
No full textWhile microarray technology has provided a versatile and high-throughput analytical tool for many research purposes, poor cross-platform assay dataset correlation has prevented the technology from finding common usage for real-world applications due to difficulties regarding the ability to validate results obtained on different platforms. Although large-scale investigations in the literature have demonstrated that cross-platform dataset correlation can be increased through the implementation of standardized interlaboratory probes, assay methodology, and analysis techniques, the degree of cross-platform concordance achievable remains significantly limited due to inherent
differences in the platforms themselves. Much of the inherent cross-platform differences limiting the extent of cross-platform dataset comparability lies with dissimilar surface properties between platforms, resulting in differential probe and target behaviors. To overcome these limitations regarding cross-platform dataset comparability, the development and use of multifunctional substrate-independent surface coatings was explored as a method to eliminate the initial differences in cross-platform surface properties and their effects on microarray performance. Specically, two types of substrate-independent surface coatings were examined: an electrostatically self-assembled polyelectrolyte multilayer and a self-polymerized polydopamine film.
The results of this investigation determined that both multifunctional substrate-independent surface coatings were capable of depositing onto a broad range of materials
and converting their surface properties into the properties of the coating itself. Additionally, when using these surface coatings as a common cross-platform interface, it
was possible to obtain highly concordant microarray datasets between platforms constructed from glass, mica, silicon, and polymer. In particular, multianalyte DNA and
protein dose-response assays performed on platforms with substrate-independent surface coatings yielded significantly higher correlation coefficients in comparison to platforms without substrate-independent surface coatings. Furthermore, it was shown how the surface properties of the multifunctional substrate-independent surface coatings can be manipulated through chemical modication in order to tailor and optimize microarray performance to suit specific applications. Utilization of substrate-independent surface coatings in such a manner can provide researchers and manufacturers with a simple, yet
effective, method to standardize microarray fabrication across different platforms while still enabling sustainable development of the technology in terms of platform material,
design, and application
Characterisation of Hardenbergia mosaic virus and development of microarrays for detecting viruses in plants
No full textA virus causing chlorosis and leaf distortion in the Western Australian endemic legume Hardenbergia comptoniana was detected by biological indexing to Chenopodium quinoa and Nicotiana benthamiana. Enzyme linked immuno-sorbent assay (ELISA) using general Potyvirus antiserum and amplification by reverse transcription polymerase chain reaction (RT-PCR) with degenerate primers indicated that it was a species of Potyvirus. It was confirmed as an unknown member of the genus Potyvirus by comparing its coat protein sequence with those of other potyviruses. The name Hardenbergia mosaic virus (HarMV) is proposed for this new virus species. Isolates of HarMV were collected from 13 sites, covering much of the natural range of its host. An experimental host range was determined using nine virus isolates tested against plants from 11 species in three families. Its infectivity on three leguminous species important in agriculture (Lupinus angustifolius, L. luteus and Trifolium subterraneum) was established.
The nucleotide (nt) sequences of the coat proteins (CP) of 28 isolates determined there was 24.1- 27.6% diversity with the closest known relative, Passion fruit woodiness virus (PWV). Studies of the nucleotide sequences of the CP showed that there was considerable intra-species divergence (mean 13.5%, maximum 20.5%) despite its relatively small geographical distribution and single known natural host. The observed broad diversity strongly suggests long genetic isolation and that HarMV evolved in the region where it was collected. An examination of its phylogeny showed that 28 isolates clustered into eight clades with high bootstrap support (6.2-20.5% inter-clade diversity). Isolates collected at locations distant to the Perth metropolitan area (Margaret River and Seabird) diverged more from isolates collected in the metropolitan area (15.4-21.1% nucleotide sequence diversity). This virus represents the first endemic species to be characterised from Western Australia.
Differences in pathogenicity and symptoms induced on key host species were seen between isolates belonging to different phylogenetic clades. Phylogenetic analysis confirmed the inclusion of HarMV within the Bean common mosaic virus group of the potyviruses and also defined a previously unreported subgroup of six previously described Potyvirus species (Clitoria virus Y, Hibbertia virus Y, PWV, Siratro 1 virus Y, and Siratro 2 virus Y), from Australia, which is further evidence for a prolonged period of genetic isolation.
Both in relation to detection of strains of HarMV, and considering the broader issues of biosecurity and parallel detection of plant viruses, a microarray based detection system was established. To optimise conditions for the development of microarrays for virus detection poly-L-lysine (PLL) coated microscope slides produced in the laboratory were compared to commercially produced PowerMatrix slides (Full Moon BioSystems). Variables tested for PLL slide production were: choice of printing buffer, probe concentration, method of immobilisation and slide blocking; and in particular the print buffer and immobilisation method had the greatest effect on the quality of PLL microarray slides. Slides printed on PLL surfaces in a high salt buffer (3x Saline sodium citrate) supplemented with 1.5M betaine and immobilised at 42oC overnight retained the highest amounts of probe DNA of the methods tested. Qualitative comparisons of the two showed more probe was retained on PowerMatrix slides which were also more reliable and consistent than the PLL slides.
Probes were designed for eight different virus species and six distinct strains of HarMV to test the potential to use microarrays to distinguish between them. Probes were designed to detect potyviruses at the genus, species and strain levels. Although there was evidence of non-specific hybridisation, the Potyvirus array was used to identify six strains of HarMV by hybridisation to species specific probes. Additionally the array was used to identify three other species of Potyvirus: Bean yellow mosaic virus, PWV and Passiflora foetida virus Y, following amplification with polyvalent PCR primers.
In further microarray tests, using labelled first strand cDNA of Potato virus X (PVX) and Potato virus Y (PVY) on an array, PVX was strongly detected in leaves known to be infected, but PVY was only weakly detected in infected leaves. Three methods of pre-amplification of virus nucleic acid before hybridisation to the array were investigated to improve the sensitivity of the assay. Two of the methods, Klenow amplification and randomly primed PCR, amplified the target virus; as confirmed by real time PCR. Of the methods tested only randomly primed PCR improved the sensitivity of the microarray. The best amplification method used genus-specific primers with adaptor sequences. This method when tested by real time PCR showed a 3.7Ct reduction for PVX and 16.8Ct for PVY. The microarray correctly identified both viruses.
In this work the first virus (HarMV) endemic to Western Australia was identified, and microarray methods were developed both to identify HarMV and other plant viruses of economic importance. The microarray approach, with further development, may be applicable as a means of identifying incursions of new viruses in a biosecurity situation
Molecular genetic methods for identifying raw materials in meat products: Diversity, opportunities and prospects
No full textIn the current economic situation, after easing the Covid pandemic restrictions, almost all laboratories, which are focused on evaluation of the conformity of food products, have faced issues in supplying for their laboratories. In this regard, in the last years many laboratories have been forced to validate new approaches and introduce new methods for assessing conformity of the food products. Very often it is not possible to use only one method to resolve the issue of the food product ingredients, especially for the purpose of traceability of their names and the used raw materials, listed on the label. Survey of the raw food materials to determine whether they correspond to the type name is a simpler task, in contrast to survey of the multicomponent food product. Many researchers have to estimate the opportunities and feasibility of application of various methodologies in their workplaces. Therefore, this review is relevant for the researchers in this field, as it focuses on aspects and special features of similar methodologies. The prospect of molecular genetic methods for identification of the raw materials used for manufacturing of meat products is presented below. This review also represents characteristics of methods for identification of the sources of raw materials used for the manufacturing of the meat products, based on the recognition of species-specific sections within the nucleic acids structures. The variety of methods (hybridization methods, polymerase chain reaction, different types of isothermal amplifications, methods using CRISPR/Cas systems), the principles of their implementation, and achieved analytical characteristics are considered. The capacities and competitive potential of various methods are discussed, as well as approaches being developed to overcome the existing limitations.In the current economic situation, after easing the Covid pandemic restrictions, almost all laboratories, which are focused on evaluation of the conformity of food products, have faced issues in supplying for their laboratories. In this regard, in the last years many laboratories have been forced to validate new approaches and introduce new methods for assessing conformity of the food products. Very often it is not possible to use only one method to resolve the issue of the food product ingredients, especially for the purpose of traceability of their names and the used raw materials, listed on the label. Survey of the raw food materials to determine whether they correspond to the type name is a simpler task, in contrast to survey of the multicomponent food product. Many researchers have to estimate the opportunities and feasibility of application of various methodologies in their workplaces. Therefore, this review is relevant for the researchers in this field, as it focuses on aspects and special features of similar methodologies. The prospect of molecular genetic methods for identification of the raw materials used for manufacturing of meat products is presented below. This review also represents characteristics of methods for identification of the sources of raw materials used for the manufacturing of the meat products, based on the recognition of species-specific sections within the nucleic acids structures. The variety of methods (hybridization methods, polymerase chain reaction, different types of isothermal amplifications, methods using CRISPR/Cas systems), the principles of their implementation, and achieved analytical characteristics are considered. The capacities and competitive potential of various methods are discussed, as well as approaches being developed to overcome the existing limitations
