Many environmental applications exist for biosensors capable of providing real-time analyses. One pressing current need is monitoring for agents of chemical- and bio-terrorism. These applications require systems that can rapidly detect small organics including nerve agents, toxic proteins, viruses, spores and whole microbes. A second area of application is monitoring for environmental pollutants. Processing of grab samples through chemical laboratories requires significant time delays in the analyses, preventing the rapid mapping and cleanup of chemical spills. The current state of development of miniaturized, integrated surface plasmon resonance (SPR) sensor elements has allowed for the development of inexpensive, portable biosensor systems capable of the simultaneous analysis of multiple analytes. Most of the detection protocols make use of antibodies immobilized on the sensor surface. The Spreeta 2000 SPR biosensor elements manufactured by Texas Instruments provide three channels for each sensor element in the system. A temperature-controlled two-element system that monitors for six analytes is currently in use, and development of an eight element sensor system capable of monitoring up to 24 different analytes will be completed in the near future. Protein toxins can be directly detected and quantified in the low picomolar range. Elimination of false positives and increased sensitivity is provided by secondary antibodies with specificity for different target epitopes, and by sensor element redundancy. Inclusion of more than a single amplification step can push the sensitivity of toxic protein detection to femtomolar levels. The same types of direct detection and amplification protocols are used to monitor for viruses and whole bacteria or spores. Special protocols are required for the detection of small molecules. Either a competition type assay where the presence of analyte inhibits the binding of antibodies to surface-immobilized analyte, or a displacement assay, where antibodies bound to analyte on the sensor surface are displaced by free analyte, can be used. The small molecule detection assays vary in sensitivity from the low micromolar range to the high picomolar.
Many environmental applications exist for biosensors capable of providing real-time analyses. One pressing current need is monitoring for agents of chemical- and bio-terrorism. These applications require systems that can rapidly detect small organics including nerve agents, toxic proteins, viruses, spores and whole microbes. A second area of application is monitoring for environmental pollutants. Processing of grab samples through chemical laboratories requires significant time delays in the analyses, preventing the rapid mapping and cleanup of chemical spills. The current state of development of miniaturized, integrated surface plasmon resonance (SPR) sensor elements has allowed for the development of inexpensive, portable biosensor systems capable of the simultaneous analysis of multiple analytes. Most of the detection protocols make use of antibodies immobilized on the sensor surface. The Spreeta 2000 SPR biosensor elements manufactured by Texas Instruments provide three channels for each sensor element in the system. A temperature-controlled two-element system that monitors for six analytes is currently in use, and development of an eight element sensor system capable of monitoring up to 24 different analytes will be completed in the near future. Protein toxins can be directly detected and quantified in the low picomolar range. Elimination of false positives and increased sensitivity is provided by secondary antibodies with specificity for different target epitopes, and by sensor element redundancy. Inclusion of more than a single amplification step can push the sensitivity of toxic protein detection to femtomolar levels. The same types of direct detection and amplification protocols are used to monitor for viruses and whole bacteria or spores. Special protocols are required for the detection of small molecules. Either a competition type assay where the presence of analyte inhibits the binding of antibodies to surface-immobilized analyte, or a displacement assay, where antibodies bound to analyte on the sensor surface are displaced by free analyte, can be used. The small molecule detection assays vary in sensitivity from the low micromolar range to the high picomolar. 相似文献
Arthropod-borne viruses (arboviruses) can persistently infect and cause limited damage to mosquito vectors. RNA interference
(RNAi) is a mosquito antiviral response important in restricting RNA virus replication and has been shown to be active against
some arboviruses. The goal of this study was to use a recombinant Sindbis virus (SINV; family Togaviridae; genus Alphavirus) that expresses B2 protein of Flock House virus (FHV; family Nodaviridae; genus Alphanodavirus), a protein that inhibits RNAi, to determine the effects of linking arbovirus infection with RNAi inhibition. 相似文献
Arthropod-borne flavivirus infection continues to cause significant morbidity and mortality worldwide. Identification of drug targets and novel antiflaviviral compounds to treat these diseases has become a global health imperative. A previous screen of 235,456 commercially available small molecules identified the 2-thioxothiazolidin-4-one family of compounds as inhibitors of the flaviviral NS5 capping enzyme, a promising target for antiviral drug development. Rational drug design methodologies enabled identification of lead compound BG-323 from this series. We have shown previously that BG-323 potently inhibits NS5 capping enzyme activity, displays antiviral effects in dengue virus replicon assays and inhibits growth of West Nile and yellow fever viruses with low cytotoxicity in vitro. In this study we further characterized BG-323’s antiviral activity in vitro and in vivo. We found that BG-323 was able to reduce replication of WNV (NY99) and Powassan viruses in culture, and we were unable to force resistance into WNV (Kunjin) in long-term culture experiments. We then evaluated the antiviral activity of BG-323 in a murine model. Mice were challenged with WNV NY99 and administered BG-323 or mock by IP inoculation immediately post challenge and twice daily thereafter. Mice were bled and viremia was quantified on day three. No significant differences in viremia were observed between BG-323-treated and control groups and clinical scores indicated both BG-323-treated and control mice developed signs of illness on approximately the same day post challenge. To determine whether differences in in vitro and in vivo efficacy were due to unfavorable pharmacokinetic properties of BG-323, we conducted a pharmacokinetic evaluation of this small molecule. Insights from pharmacokinetic studies indicate that BG-323 is cell permeable, has a low efflux ratio and does not significantly inhibit two common cytochrome P450 (CYP P450) isoforms thus suggesting this molecule may be less likely to cause adverse drug interactions. However, the T1/2 of BG-323 was suboptimal and the percent of drug bound to plasma binding proteins was high. Future studies with BG-323 will be aimed at increasing the T1/2 and determining strategies for mitigating the effects of high plasma protein binding, which likely contribute to low in vivo efficacy. 相似文献
Two different serotypes of the genus Nitrosomonas were isolated from samples of the sewage plant Heidelberg. These nitrifiers were enumerated in activated sludge of various other sewage plants after immunofluorescent labeling and staining with propidium iodide by flow cytometry. The concentrations of these serotypes of Nitrosomonas spp. were in the range of 0.1 to 2%. Also, a test for the determination of the activity of ammonia-oxidizing bacteria was developed. Nitrite-oxidizing bacteria were specifically inhibited with sodium chlorate, and the activity of ammonia-oxidizing bacteria could be calculated from the increase of nitrite. Concentrations and activities of ammonia oxidizers were measured for a period of 6 months in the sewage plant Heidelberg. With one exception, activities and concentrations of ammonia-oxidizing bacteria decreased and increased in parallel. 相似文献
The evolutionary origin of the pinnipeds (seals, sea lions, and walruses)
is still uncertain. Most authors support a hypothesis of a monophyletic
origin of the pinnipeds from a caniform carnivore. A minority view suggests
a diphyletic origin with true seals being related to the mustelids (otters
and ferrets). The phylogenetic relationships of the walrus to other
pinniped and carnivore families are also still particularly problematic.
Here we examined the relative support for mono- and diphyletic hypotheses
using DNA sequence data from the mitochondrial small subunit (12S) rRNA and
cytochrome b genes. We first analyzed a small group of taxa representing
the three pinniped families (Phocidae, Otariidae, and Odobenidae) and
caniform carnivore families thought to be related to them. We inferred
phylogenetic reconstructions from DNA sequence data using standard
parsimony and neighbor-joining algorithms for phylogenetic inference as
well as a new method called spectral analysis (Hendy and Penny) in which
phylogenetic information is displayed independently of any selected tree.
We identified and compensated for potential sources of error known to lead
to selection of incorrect phylogenetic trees. These include sampling error,
unequal evolutionary rates on lineages, unequal nucleotide composition
among lineages, unequal rates of change at different sites, and
inappropriate tree selection criteria. To correct for these errors, we
performed additional transformations of the observed substitution patterns
in the sequence data, applied more stringent structural constraints to the
analyses, and included several additional taxa to help resolve long,
unbranched lineages in the tree. We find that there is strong support for a
monophyletic origin of the pinnipeds from within the caniform carnivores,
close to the bear/raccoon/panda radiation. Evidence for a diphyletic origin
was very weak and can be partially attributed to unequal nucleotide
compositions among the taxa analyzed. Subsequently, there is slightly more
evidence for grouping the walrus with the eared seals versus the true
seals. A more conservative interpretation, however, is that the walrus is
an early, but not the first, independent divergence from the common
pinniped ancestor.
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