Sensitive CH4 detection applying quantum cascade laser based optical feedback cavity-enhanced absorption spectroscopy.

We report on sensitive detection of atmospheric methane employing quantum cascade laser based optical feedback cavity-enhanced absorption spectroscopy (OF-CEAS). An instrument has been built utilizing a continuous-wave distributed feedback quantum cascade laser (cw-QCL) with a V-shaped cavity, a common arrangement that reduces feedback to the laser from non-resonant reflections. The spectrometer has a noise equivalent absorption coefficient of 3.6 × 10-9 cm-1 Hz-1/2 for a spectral scan of CH4 at 7.39 µm. From an Allan-Werle analysis a detection limit of 39 parts per trillion of CH4 at atmospheric pressure within 50 s acquisition time was found.

[Diagnostic value of the quantitative HBsAg determination in hepatitis B infections]. / Diagnostischer Wert der quantitativen HBsAg-Bestimmung bei der Hepatitis B-Infektion.

Introduction of systematic hepatitis B vaccination has lead to a strong decrease of new infections, but there are still a high numbers of chronically infected persons suffering on long-term complications. Using quantitative assays for the determination of HbsAg (qHBsAg) has improved our understanding of chronic hepatitis B (CHB). The concentrations of HBsAg are strongly varying through the different stages of infection. The quantitative determination of HBsAg does not only yield in additional information to the infection activity, but also provides data for an improved follow up independent from the virus load. As to the prediction of disease progression, low-viremic carriers with high HbsAg levels have been shown to be at higher risk of HBeAg negative hepatitis, cirrhosis and hepatocellular carcinoma (HCC). Although, quantitative HBsAg determination has been widely used in CHB patients receiving pegylated interferon therapy, the HbsAg decline is slow compared to HBV-DNA levels during nucleos(t)ide analogue (NUC) therapy. However a rapid HbsAg decline during NUC therapy may identify patients who will finally clear HbsAg. A 6- to 12-monthly assessment of HbsAg level could be considered during NUC therapy. Taking these lines of evidence together, qHBsAg can complement HBV-DNA levels to optimize the management of CHB patients.

[Hepatitis C diagnostics: clinical evaluation of the HCV-core antigen determination]. / Hepatitis-C-Diagnostik: Klinische Evaluierung der HCV-Core-Antigenbestimmung.

BACKGROUND: The aim of the evaluation was to investigate the relevance of the HCV-core antigen testing for the diagnosis and monitoring of HCV infections in the daily routine. Up to now, most of the serological diagnostics was performed as determination of antibodies while the determination of activity and the monitoring of antiviral therapy were checked by HCV RNA PCR. METHODS: The routine requests for HCV-core antigen of a private laboratory were analyzed for a period of two years. RESULTS: The determination of HCV antigen highly correlates with the quantitative measurement of HCV RNA (r = 0.73), p = 0.0003). The diagnostic window is comparable with that of the HCV PCR (27.1 ±â€Š12.8 d vs. 23.9 ±â€Š9.2 d, p = 0.11). The sensitivity of the HCV antigen assay was 99.0 % with a specificity of 99.2 %. 54.3 % of the confirmed antibody positive samples were also antigen positive. Only in 3 of 560 HCV-RNA positive samples HCV antigen was not detectable, but 3 samples without HCV antibodies were confirmed positive for HCV antigen. CONCLUSIONS: The HCV antigen assay is a suitable tool for the detection of chronic active HCV infections, for the early diagnosis of acute infections and for testing of HCV in patient with immunodeficiency. The HCV antigen assay valuable completion of serological testing for HCV.

Performance evaluation of a new fourth-generation HIV combination antigen-antibody assay.

Education and diagnostic tests capable of early detection represent our most effective means of preventing transmission of human immunodeficiency virus (HIV). The importance of early detection is underlined by studies demonstrating increased life expectancy following early initiation of antiviral treatment. The Elecsys(®) HIV combi PT assay is a fourth-generation antigen-antibody combination assay developed to allow earlier detection of seroconversion, and to have increased sensitivity and improved specificity. We aimed to determine how early the assay could detect infection compared with existing assays; whether all HIV variants could be detected; and the assay's specificity using samples from blood donors, routine specimens, and patients with potential cross-reacting factors. Samples were identified as positive by the Elecsys(®) assay 4.9 days after a positive polymerase chain reaction result (as determined by the panel supplier), which was earlier than the 5.3-7.1 days observed with comparators. The analytical sensitivity of the Elecsys(®) HIV combi PT assay for the HIV-1 p24 antigen was 1.05 IU/mL, which compares favorably with the comparator assays. In addition, the Elecsys(®) assay identified all screened HIV subtypes and displayed greater sensitivity to HIV-2 homologous antigen and antibodies to HIV-1 E and O and HIV-2 than the other assays. Overall, the specificity of the Elecsys(®) assay was 99.88 % using samples from blood donors and 99.81 % when analyzing unselected samples. Potential cross-reacting factors did not interfere with assay performance. The Elecsys(®) HIV combi PT assay is a sensitive and specific assay that has been granted the CE mark according to Directive 2009/886/EC.

Clinical evaluation of new automated cytomegalovirus IgM and IgG assays for the Elecsys(®) analyser platform.

Cytomegalovirus (CMV) is a leading cause of physical and neurological abnormalities in newborns. Hence, the diagnosis of CMV infection in pregnant women is necessary in order to allow appropriate management of their pregnancy. New assays have been developed for the Roche Elecsys® immunoassay platform that detect CMV-specific immunoglobulin (Ig)M and IgG, with the IgM assay designed to target IgM produced at the start of infection rather than IgM persisting later in infection. This study aimed to evaluate the performance of the new assays compared with other commercial kits widely distributed in laboratories. The performance of the Elecsys and comparator CMV IgM and IgG assays was assessed using 967 preselected patient samples characterised by CMV infection status, as well as being compared using 1,668 unselected clinical samples. The Elecsys CMV IgM and IgG assays performed consistently with comparator assays using the preselected samples. The Elecsys CMV IgM assay showed improved sensitivity compared with the Enzygnost® assay in primary infection (91.2 % vs. 79.4 %) and improved specificity over the Architect® assay in potentially cross-reacting samples (94.1 % vs. 82.4 %). The Elecsys IgM assay reported fewer positive results in the later stages of CMV infection compared with ETI-CYTOK-M ELISA, while the Elecsys IgG assay reported slightly fewer negative results in the early stages of infection compared with ETI-CYTOK-G ELISA. There was good agreement between Elecsys and comparator assays using unselected clinical samples (range 90.4-99.4 %). The Elecsys CMV IgM and IgG assays compare well with routinely used assays and are suitable for clinical use.

Reliability of high-density surface electromyography for assessing characteristics of the thoracic erector spinae during static and dynamic tasks.

PURPOSE: To establish intra- and inter-session reliability of high-density surface electromyography (HDEMG)-derived parameters from the thoracic erector spinae (ES) during static and dynamic goal-directed voluntary movements of the trunk, and during functional reaching tasks. METHODS: Twenty participants performed: 1) static trunk extension, 2) dynamic trunk forward and lateral flexion, and 3) multidirectional functional reaching tasks on two occasions separated by 7.5 ± 1.2 days. Muscle activity was recorded bilaterally from the thoracic ES. Root mean square (RMS), coordinates of the barycentre, mean frequency (MNF), and entropy were derived from the HDEMG signals. Reliability was determined with intraclass correlation coefficient (ICC), coefficient of variation, and standard error of measurement. RESULTS: Good-to-excellent intra-session reliability was found for all parameters and tasks (ICC: 0.79-0.99), whereas inter-session reliability varied across tasks. Static tasks demonstrated higher reliability in most parameters compared to functional and dynamic tasks. Absolute RMS and MNF showed the highest overall reliability across tasks (ICC: 0.66-0.98), while reliability of the barycentre was influenced by the direction of the movements. CONCLUSION: RMS and MNF derived from HDEMG show consistent inter-session reliability in goal-directed voluntary movements of the trunk and reaching tasks, whereas the measures of the barycentre and entropy demonstrate task-dependent reliability.

Rapid passage signals from a vibrationally excited target molecule: a pump and probe experiment with continuous wave quantum cascade lasers.

Two 5 µm continuous wave quantum cascade lasers are used to perform a counterpropagating pump and probe experiment on a low pressure sample of nitric oxide. The strong pump field excites a fundamental rovibrational transition and the weaker probe field is tuned to the corresponding rotationally resolved hot band transition. When both light fields are in resonance, rapid passage is observed in the hot band absorption lineshape arising from a minimally damped and velocity-selected sample of molecules in the v=1 state. The measured rapid passage signals are well described by a two-level model based on the optical Bloch equations.

Multicenter evaluation of a fully automated third-generation anti-HCV antibody screening test with excellent sensitivity and specificity.

Early detection of hepatitis C virus (HCV) is an important step in preventing progression to cirrhosis and hepatocellular carcinoma. Serologic assays for anti-hepatitis C (anti-HCV) antibody are valuable first-line tests in the screening and diagnosis of HCV infection. The aim of this multicenter study was to compare the Elecsys(®) Anti-HCV assay with alternative CE-marked Anti-HCV antibody assays against a range of samples that included 1,138 blood donors, 3,553 unselected routine daily specimens, and 46 pre-selected seroconversion panels. Specificity of the Elecsys Anti-HCV assay was 99.5% with blood donor samples and 99.4% with routine clinical specimens. These were similar to those obtained with the Prism(®) Anti-HCV, Architect(®) Anti-HCV assay, ADVIA(®) Centaur Anti-HCV assay and Vitros(®) Eci aHCV assays. Seroconversion sensitivity for the Elecsys Anti-HCV assay was similar to that of the Architect Anti-HCV, AxSYM HCV version 3.0, ADVIA Centaur Anti-HCV, and Vitros Eci aHCV assays. In fact, seroconversion testing on 46 commercially available panels showed that the difference in first detecting a positive blood sample was less than one day between assays (not statistically significant). The Elecsys Anti-HCV assay as well as the Architect, Prism, and Vitros Anti-HCV immunoassays revealed a seroconversion sensitivity of 100%, whereas the ADVIA Centaur HCV immunoassay showed a sensitivity of only 97.5% (39/40). Overall, the performance of the Elecsys Anti-HCV assay was similar to the performances of the comparator CE-marked Anti-HCV antibody assays.

Rapid passage signals induced by chirped quantum cascade laser radiation: K state dependent-delay effects in the nu2 band of NH3.

In this Letter, a 10 microm quantum cascade laser operating in the intrapulse mode is used observe rapid passage (RP) effects within a 40 cm single-pass gas cell containing low pressures of NH(3). The laser tuning range allows the rotational states J=2 with K=0, 1, and 2 to be probed. We show that the RP structures change as a function of optical density and that the magnitude of the delay in the switch from absorption to emission as a function of increased gas pressure is dependent upon the initial value of K. These measurements are qualitatively well modeled using the Maxwell-Bloch equations.

Characterization of an external cavity diode laser based ring cavity NICE-OHMS system.

The performance of an external cavity diode laser based noise immune cavity enhanced optical heterodyne molecular spectrometer is presented. To reduce the noise on the signal a ring cavity and a circuit to remove residual amplitude modulation on the pre-cavity laser radiation was implemented. We demonstrate a sensitivity of 4 x 10(-11) cm(-1) Hz(-1/2) using a cavity with a finesse of 2600 on a Doppler-broadened transition of CH(4) at 6610.063 cm(-1).

CYGD: the Comprehensive Yeast Genome Database.

The Comprehensive Yeast Genome Database (CYGD) compiles a comprehensive data resource for information on the cellular functions of the yeast Saccharomyces cerevisiae and related species, chosen as the best understood model organism for eukaryotes. The database serves as a common resource generated by a European consortium, going beyond the provision of sequence information and functional annotations on individual genes and proteins. In addition, it provides information on the physical and functional interactions among proteins as well as other genetic elements. These cellular networks include metabolic and regulatory pathways, signal transduction and transport processes as well as co-regulated gene clusters. As more yeast genomes are published, their annotation becomes greatly facilitated using S.cerevisiae as a reference. CYGD provides a way of exploring related genomes with the aid of the S.cerevisiae genome as a backbone and SIMAP, the Similarity Matrix of Proteins. The comprehensive resource is available under http://mips.gsf.de/genre/proj/yeast/.

Discovering regulatory elements in non-coding sequences by analysis of spaced dyads.

The application of microarray and related technologies is currently generating a systematic catalog of the transcriptional response of any single gene to a multiplicity of experimental conditions. Clustering genes according to the similarity of their transcriptional response provides a direct hint to the regulons of the different transcription factors, many of which have still not been characterized. We have developed a new method for deciphering the mechanism underlying the common transcriptional response of a set of genes, i.e. discovering cis -acting regulatory elements from a set of unaligned upstream sequences. This method, called dyad analysis, is based on the observation that many regulatory sites consist of a pair of highly conserved trinucleotides, spaced by a non-conserved region of fixed width. The approach is to count the number of occurrences of each possible spaced pair of trinucleotides, and to assess its statistical significance. The method is highly efficient in the detection of sites bound by C(6)Zn(2)binuclear cluster proteins, as well as other transcription factors. In addition, we show that the dyad and single-word analyses are efficient for the detection of regulatory patterns in gene clusters from DNA chip experiments. In combination, these programs should provide a fast and efficient way to discover new regulatory sites for as yet unknown transcription factors.

Statistical analysis of yeast genomic downstream sequences reveals putative polyadenylation signals.

The study of a few genes has permitted the identification of three elements that constitute a yeast polyadenyl-ation signal: the efficiency element (EE), the positioning element and the actual site for cleavage and poly-adenyl-ation. In this paper we perform an analysis of oligonucleotide composition on the sequences located downstream of the stop codon of all yeast genes. Several oligonucleotide families appear over-represented with a high significance (referred to herein as 'words'). The family with the highest over-representation includes the oligonucleotides shown experimentally to play a role as EEs. The word with the highest score is TATATA, followed, among others, by a series of single-nucleotide variants (TATGTA, TACATA, TAAATA.) and one-letter shifts (ATATAT). A position analysis reveals that those words have a high preference to be in 3' flanks of yeast genes and there they have a very uneven distribution, with a marked peak around 35 bp after the stop codon. Of the predicted ORFs, 85% show one or more of those sequences. Similar results were obtained using a data set of EST sequences. Other clusters of over-represented words are also detected, namely T- and A-rich signals. Using these results and previously known data we propose a general model for the 3' trailers of yeast mRNAs.

Extracting regulatory sites from the upstream region of yeast genes by computational analysis of oligonucleotide frequencies.

We present here a simple and fast method allowing the isolation of DNA binding sites for transcription factors from families of coregulated genes, with results illustrated in Saccharomyces cerevisiae. Although conceptually simple, the algorithm proved efficient for extracting, from most of the yeast regulatory families analyzed, the upstream regulatory sequences which had been previously found by experimental analysis. Furthermore, putative new regulatory sites are predicted within upstream regions of several regulons. The method is based on the detection of over-represented oligonucleotides. A specificity of this approach is to define the statistical significance of a site based on tables of oligonucleotide frequencies observed in all non-coding sequences from the yeast genome. In contrast with heuristic methods, this oligonucleotide analysis is rigorous and exhaustive. Its range of detection is however limited to relatively simple patterns: short motifs with a highly conserved core. These features seem to be shared by a good number of regulatory sites in yeast. This, and similar methods, should be increasingly required to identify unknown regulatory elements within the numerous new coregulated families resulting from measurements of gene expression levels at the genomic scale. All tools described here are available on the web at the site http://copan.cifn.unam.mx/Computational_Biology/ yeast-tools

Interactive visualization and exploration of relationships between biological objects.

Genome sequencing and microarray technology produce ever-increasing amounts of complex data that need analysis. Visualization is an effective analytical technique that exploits the ability of the human brain to process large amounts of data. Here, we review traditional visualization methods based on clustering and tree representation, and also describe an alternative approach that involves projecting objects onto a Euclidean space in a way that reflects their structural or functional distances. Data are visualized without preclustering and can be dynamically explored by the user using 'virtual-reality'. We illustrate this approach with two case studies from protein topology and gene expression.

Establishement of the dorso-ventral pattern during embryonic development of drosophila melanogasater: a logical analysis

This report focuses on dorso-ventral patterning in the segmented region of the Drosophila melanogaster embryo. According to the concept of positional information, this pattern results from the different response of cells to the Dorsal-protein morphogen. This protein shows a distribution gradient along the dorso-ventral axis, with the highest concentration on the ventral side. Using the generalized logical formalism developed by R. Thomas and co-workers, the different cellular responses were analysed in terms of the intracellular loops between the regulatory genes. Two positive loops were found to be involved, each constituting a switch which can be acted upon by the Dorsal morphogen to determine the different cell types that make up the embryonic dorso-ventral pattern. The novelty in this use of generalized logical formalism is the employment of a multilevel variable to represent a morphogen gradient. The proposed model accounts for the essential qualitative effects of the Dorsal gradient in the dorso-ventral determination process. Three main conclusions may be drawn. Firstly, the gene twist needs to have two functional threshold concentrations, one for autoactivation and the other for activation of the gene snail. Secondly, the autoactivation threshold must be smaller than that which activates snail. Thirdly, the action of the gene snail on the maintenance function of the gene twist is crucial for cells to be able to choose between the mesoderm or neuroectoderm developmental pathways. Furthermore, it is predicted that if the gene snail shows autoregulation, this will not be crucial for the determination of the embryonic D-V pattern. Copyright 1997 Academic Press Limited Copyright 1997 Academic Press Limited

Production mechanisms of NH and NH2 radicals in N2-H2 plasmas.

We measured the densities of NH and NH(2) radicals by cavity ring-down spectroscopy in N(2)-H(2) plasmas expanding from a remote thermal plasma source and in N(2) plasmas to which H(2) was added in the background. The NH radical was observed via transitions in the (0,0), (1,1), and (2,2) vibrational bands of the A(3)Pi <-- X(3)Sigma- electronic transition and the NH(2) radical via transitions in the (0,9,0) <-- (0,0,0) band of the A(2)A(1) <-- X(2)B(1) electronic transition. The measurements revealed typical densities of 5 x 10(18) m(-3) for the NH radical in both plasmas and up to 7 x 10(18) m(-3) for the NH(2) radical when N(2) and H(2) are both fed through the plasma source. In N(2) plasma with H(2) injected in the background, no NH(2) was detected, indicating that the density is below our detection limit of 3 x 1016 m-3. The error in the measured densities is estimated to be around 20%. From the trends of the NH(x) radicals as a function of the relative H(2) flow to the total N(2) and H(2) flow at several positions in the expanding plasma beam, the key reactions for the formation of NH and NH(2) have been determined. The NH radicals are mainly produced via the reaction of N atoms emitted by the plasma source with H(2) molecules with a minor contribution from the reaction of N+ with H(2). The NH(2) radicals are formed by reactions of NH(3) molecules, produced at the walls of the plasma reactor, and H atoms emitted by the plasma source. The NH radicals can also be produced by H abstraction of NH(2) radicals. The flux densities of the NH(x) radicals with respect to the atomic radicals are appreciable in the first part of the expansion. Further downstream the NH(x) radicals are dissociated, and their densities become smaller than those of the atomic radicals. It is concluded that the NH(x) radicals play an important role as precursors for the N and H atoms, which are key to the surface production of N(2), H(2), and NH(3) molecules.

A web site for the computational analysis of yeast regulatory sequences.

A series of computer programs were developed for the analysis of regulatory sequences, with a special focus on yeast. These tools are publicly available on the web (http://copan.cifn.unam. mx/Computational_Biology/yeast-tools or http://www.ucmb.ulb.ac. be/bioinformatics/rsa-tools/). Basically, three classical problems can be addressed: (a) search for known regulatory patterns in the upstream regions of known genes; (b) discovery of unknown regulatory patterns within a set of upstream regions known to be co-regulated; (c) search for unknown genes potentially regulated by a known transcription factor. Each of these tasks can be performed on basis of a simple (string) or more refined (matrix) description of the regulatory patterns. A feature-map program automatically generates visual representations of the positions at which patterns were found. The site also provides a series of general utilities, such as generation of random sequence, automatic drawing of XY graphs, interconversions between sequence formats, etc. Several tools are linked together to allow their sequential utilization (piping), but each one can also be used independently by filling the web form with external data. This widens the scope of the site to the analysis of non-regulatory and/or non-yeast sequences.

Combining pattern discovery and discriminant analysis to predict gene co-regulation.

MOTIVATION: Several pattern discovery methods have been proposed to detect over-represented motifs in upstream sequences of co-regulated genes, and are for example used to predict cis-acting elements from clusters of co-expressed genes. The clusters to be analyzed are often noisy, containing a mixture of co-regulated and non-co-regulated genes. We propose a method to discriminate co-regulated from non-co-regulated genes on the basis of counts of pattern occurrences in their non-coding sequences. METHODS: String-based pattern discovery is combined with discriminant analysis to classify genes on the basis of putative regulatory motifs. RESULTS: The approach is evaluated by comparing the significance of patterns detected in annotated regulons (positive control), random gene selections (negative control) and high-throughput regulons (noisy data) from the yeast Saccharomyces cerevisiae. The classification is evaluated on the annotated regulons, and the robustness and rejection power is assessed with mixtures of co-regulated and random genes.