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PubMed2005

Inter-laboratory validation of PCR-based detection of Mycobacterium tuberculosis in formalin-fixed, paraffin-embedded tissues.

The present study is based on the initiative for quality assurance in pathology of the German Society of Pathology and the Professional Association of German Pathologists. Four panel laboratories with experience and expertise in polymerase chain reaction (PCR) detection of Mycobacterium tuberculosis were selected to establish the prerequisites for continuous external laboratory trials, in particular, by providing pre-tested specimens and evaluation criteria for participating institutes. In the first step, the four panel laboratories performed an internal trial to test their own reliability and reproducibility. Paraffin sections and DNA preparations from 34 tissues (25 clinical specimens and 9 controls) totalling to 66 samples were evaluated by each panel institute according to their own protocols. The methodologies differed and are described in detail. Despite these differences, a high degree of inter-laboratory reliability was achieved. In this report, we summarise our results including the correlation with the histology and provide recommendations for applying PCR-based methodology for the detection of mycobacterial DNA in surgical specimens. Supplementary data are available online at http://www.charite.de/ch/patho (rubric "Forschung"). Pre-tested specimens are now available for the external trial and can be ordered from the steering institute via Oligene (http://www.oligene.com/). All molecular pathology laboratories are invited to participate in this quality assurance initiative.

Animals↗
PubMed1989

Prevalence of very severe headache in a large US metropolitan area.

Reliable data regarding prevalence of headache, in general, and migraine, in particular, remain elusive. Methodological inconsistencies and inadequacies are largely responsible. The present study uses a random sampling technique to assess the prevalence of very severe headache. Estimates of headache prevalence were obtained for two time periods: indeterminate/lifetime and the past three months. The resulting prevalence estimates were 20.4 and 15.2%, respectively, for women, and 11.2 and 8.8%, respectively, for men. The estimates are compared with those of previous studies. The present study is compared with these previous studies and suggestions are offered for future research. Aspects of the treatment experience of respondents with poorly controlled headache are also addressed.

Adult↗
PubMed1997

Modeling protein stability: a theoretical analysis of the stability of T4 lysozyme mutants.

Free energy calculations were conducted to determine the relative stability of the unnatural amino acid mutants of T4 lysozyme norvaline (Nvl) and O-methyl-serine (Mse) and of alanine at residue 133, which is leucine in the native sequence. These calculations were performed both to assess the validity of the methodology and to gain a better understanding of the forces which contribute to protein stability. Peptides of different length were used to model the denatured state. Restraints were employed to force sampling of the side chain chi1 dihedral of the perturbed side chain, and the effect of protein repacking in response to mutation was studied through the use of different constraint sets. In addition, the convergence behavior and hysteresis of the simulations in the folded and unfolded states were determined. The calculated results agree well with experiment, + 1.84 versus + 1.56 kcal/mol for Mse-->Nvl and -3.48 versus -2.2 to -3.6 kcal/mol for Nvl-->Ala. We find that free energy calculations can provide useful insights to protein stability when conducted carefully on a well chosen system. Our results suggest that loss of packing interactions in the native state is a major source of destabilization for mutants which decrease the amount of buried nonpolar surface area and that subtle responses of the backbone affect the magnitude of the loss of stability. We show that the conformational freedom of the chi1 dihedral has a noticeable effect on protein stability and that the solvation of amino acid side chains is strongly influenced by interactions with the peptide backbone.

Amino Acid Sequence↗
PubMed2001

An efficient and robust statistical modeling approach to discover differentially expressed genes using genomic expression profiles.

We have developed a statistical regression modeling approach to discover genes that are differentially expressed between two predefined sample groups in DNA microarray experiments. Our model is based on well-defined assumptions, uses rigorous and well-characterized statistical measures, and accounts for the heterogeneity and genomic complexity of the data. In contrast to cluster analysis, which attempts to define groups of genes and/or samples that share common overall expression profiles, our modeling approach uses known sample group membership to focus on expression profiles of individual genes in a sensitive and robust manner. Further, this approach can be used to test statistical hypotheses about gene expression. To demonstrate this methodology, we compared the expression profiles of 11 acute myeloid leukemia (AML) and 27 acute lymphoblastic leukemia (ALL) samples from a previous study (Golub et al. 1999) and found 141 genes differentially expressed between AML and ALL with a 1% significance at the genomic level. Using this modeling approach to compare different sample groups within the AML samples, we identified a group of genes whose expression profiles correlated with that of thrombopoietin and found that genes whose expression associated with AML treatment outcome lie in recurrent chromosomal locations. Our results are compared with those obtained using t-tests or Wilcoxon rank sum statistics.

Acute Disease↗
PubMed2005

Changes in mental health literacy about depression: South Australia, 1998 to 2004.

OBJECTIVE: To identify changes in mental health literacy in regard to depression between 1998 and 2004. DESIGN AND SETTING: Face-to-face interviews with a random and representative sample of the South Australian population in 2004, compared with a similarly conducted survey in 1998 that used the same vignette, questions and methodology. PARTICIPANTS: 3015 randomly selected participants, aged 15 years and over. MAIN OUTCOME MEASURES: Responses to both open-ended and direct questions about symptoms and treatment options for depression. RESULTS: The 3015 interviews conducted represented a response rate of 65.9%. Compared with 1998, in 2004 there was a significant increase in the proportion of people recognising depression in the vignette, acknowledging personal experience of depression, and perceiving professional assistance to be more helpful and less harmful. However, although more people nominated psychiatrists or psychologists as therapists of choice, the difference between 1998 and 2004 was not significant. CONCLUSIONS: There has been a significant increase in mental health literacy, at least as regards depression, in the South Australian community between 1998 and 2004. The lack of significant change in psychiatrists and/or psychologists being perceived as therapists of choice is of concern and suggests that community education about their expertise may be appropriate.

Adolescent↗
PubMed1984

A method for eliciting beliefs about food and child feeding in Papua New Guinea: the MACHIK Interview.

A large proportion of Papua New Guinean children exhibit growth retardation early in the first year of life despite continued breastfeeding. Numerous investigators have suggested that traditional concepts and practices concerned with the introduction of solid foods are responsible for the observed poor growth. While there is recently some evidence to support this idea in one coastal population, the hypothesis warrants empirical testing. It is suggested that food taboos and other concepts and behaviours related to feeding children be carefully described and, if possible, quantified as a function of age, thereby linking behaviour to growth curves in a testable manner. A directed interview, derived from extended ethnographic research in community foodways and known as the Maternal and Child Health Indigenous Knowledge, or MACHIK Interview has been developed for use in Papua New Guinea societies. In this paper a short form designed specifically for gathering information on beliefs about child feeding is presented. Annotations to the questions are included to enable use by those having little experience or training in ethnographic methods.

Adult↗
PubMed1982

The adolescent boy and girl: first and other early experiences with intercourse from a representative sample of Swedish school adolescents.

Attitudes and sexual experiences among adolescents have been studied using data from a representative sample of pupils in tenth grade in Uppsala, the fourth largest city in Sweden. Sex differences in coital experience, coital frequency, and evaluation of coital frequency are analyzed. The importance of traditional sex roles is discussed, and the differential sexual socialization of boys and girls is commented on. The data help convey a picture of an adolescent boy who feels deprived of heterosexual outlets and whose sexual relations with girls are sudden, unplanned, and characterized by limited emotional involvement; the adolescent boy is also quite discontent. This picture is contrasted with that of the girls, about whom it is found that, compared with the boys, a greater proportion have coital experience, the average age at first intercourse is lower, and the average number of experiences with intercourse during the last month is higher; to a greater extent than the boys, the girls are satisfied with how often they have intercourse.

Adolescent↗
PubMed2006

Dephased MRI.

In this work gradient dephasing is treated as a mechanism for manipulating contrast in otherwise conventional MR images. The paper provides a theoretical and experimental framework for this approach. It starts from the observation that dephasing gradients invoke a shift in k-space. From this it is inferred that the effects of in-plane and through-plane dephasing can be systematically explored in the context of any given imaging experiment by sampling k-space more widely and densely than dictated by the field of view (FOV) and the spatial resolution of the desired images. The oversampled k-space allows an ensemble of lower-resolution dephased images to be reconstructed in which the degree and direction of dephasing are determined by the off-center position of the reconstruction window. The efficacy of this approach is demonstrated for standard gradient-echo acquisitions in a phantom. The results indicate the potential of the proposed methodology for evaluating 3D image data and optimizing gradient dephasing in applications that rely on the exploitation of partial volume and susceptibility effects (e.g., tracking interventional devices and tracing magnetically labeled substances).

Gels↗
PubMed2000

Developing a model of collaborative research: the complexities and challenges of implementation.

While the benefits of collaborative research have been well documented, fewer publications exist regarding the complex, problematic issues involved in these undertakings. This paper offers an integrated collaborative research model to depict the complexities and challenges of initiation and implementation of a 2 1/2 year joint research project between a community hospital and a university school of nursing in Southern Ontario, Canada. A sampling of the experiences of the researchers are analyzed to clarify the dynamic and often competing issues and interactions involved in encouraging hospital-wide research involvement during periods of organizational growth and change. The model reflects the simultaneous interaction of organizational, change and collaborative processes while maintaining the rigor of the research (RCT), and ensuring minimal disruption to the service agency. Quantitative outcomes of this collaboration are presented through an analysis of participant involvement on multiple organizational levels. Recommendations for future collaborative research, including design and methodological issues and collaborative and change strategies are offered. The complexity of balancing the necessary trade-offs required of successful collaborative research are highlighted and will be useful to those considering and planning future collaborations.

Clinical Nursing Research↗
PubMed2003

NMR studies of the interaction of a type II dihydrofolate reductase with pyridine nucleotides reveal unexpected phosphatase and reductase activity.

The interaction of type II R67 dihydrofolate reductase (DHFR) with its cofactor nicotinamide adenine dinucleotide phosphate (NADP(+)) has been studied using nuclear magnetic resonance (NMR). Doubly labeled [U-(13)C,(15)N]DHFR was obtained from Escherichia coli grown on a medium containing [U-(13)C]-D-glucose and (15)NH(4)Cl, and the 16 disordered N-terminal amino acids were removed by treatment with chymotrypsin. Backbone and side chain NMR assignments were made using triple-resonance experiments. The degeneracy of the amide (1)H and (15)N shifts of the tetrameric DHFR was preserved upon addition of NADP(+), consistent with kinetic averaging among equivalent binding sites. Analysis of the more titration-sensitive DHFR amide resonances as a function of added NADP(+) gave a K(D) of 131 +/- 50 microM, consistent with previous determinations using other methodology. We have found that the (1)H spectrum of NADP(+) in the presence of the R67 DHFR changes as a function of time. Comparison with standard samples and mass spectrometric analysis indicates a slow conversion of NADP(+) to NAD(+), i.e., an apparent NADP(+) phosphatase activity. Studies of this activity in the presence of folate and a folate analogue support the conclusion that this activity results from an interaction with the DHFR rather than a contaminating phosphatase. (1)H NMR studies of a mixture of NADP(+) and NADPH in the presence of the enzyme reveal that a ternary complex forms in which the N-4A and N-4B nuclei of the NADPH are in the proximity of the N-4 and N-5 nuclei of NADP(+). Studies using the NADP(+) analogue acetylpyridine adenosine dinucleotide phosphate (APADP(+)) demonstrated a low level of enzyme-catalyzed hydride transfer from NADPH. Analysis of DHFR backbone dynamics revealed little change upon binding of NADP(+). These additional catalytic activities and dynamic behavior are in marked contrast to those of type I DHFR.

Amides↗
PubMed2003

Preventing central venous catheter-associated primary bloodstream infections: characteristics of practices among hospitals participating in the Evaluation of Processes and Indicators in Infection Control (EPIC) study.

OBJECTIVES: To describe the conceptual framework and methodology of the Evaluation of Processes and Indicators in Infection Control (EPIC) study and present results of CVC insertion characteristics and organizational practices for preventing BSIs. The goal of the EPIC study was to evaluate relationships among processes of care, organizational characteristics, and the outcome of BSI. DESIGN: This was a multicenter prospective observational study of variation in hospital practices related to preventing CVC-associated BSIs. Process of care information (eg, barrier use during insertions and experience of the inserting practitioner) was collected for a random sample of approximately 5 CVC insertions per month per hospital during November 1998 to December 1999. Organization demographic and practice information (eg, surveillance activities and staff and ICU nurse staffing levels) was also collected. SETTING: Medical, surgical, or medical-surgical ICUs from 55 hospitals (41 U.S. and 14 international sites). PARTICIPANTS: Process information was obtained for 3,320 CVC insertions with an average of 58.2 (+/- 16.1) insertions per hospital. Fifty-four hospitals provided policy and practice information. RESULTS: Staff spent an average of 13 hours per week in study ICU surveillance. Most patients received nontunneled, multiple lumen CVCs, of which fewer than 25% were coated with antimicrobial material. Regarding barriers, most clinicians wore masks (81.5%) and gowns (76.8%); 58.1% used large drapes. Few hospitals (18.1%) used an intravenous team to manage ICU CVCs. CONCLUSIONS: Substantial variation exists in CVC insertion practice and BSI prevention activities. Understanding which practices have the greatest impact on BSI rates can help hospitals better target improvement interventions.

Catheterization, Central Venous↗
PubMed2006

Possible stimuli for strength and power adaptation : acute metabolic responses.

The metabolic response to resistance exercise, in particular lactic acid or lactate, has a marked influence upon the muscular environment, which may enhance the training stimulus (e.g. motor unit activation, hormones or muscle damage) and thereby contribute to strength and power adaptation. Hypertrophy schemes have resulted in greater lactate responses (%) than neuronal and dynamic power schemes, suggesting possible metabolic-mediated changes in muscle growth. Factors such as age, sex, training experience and nutrition may also influence the lactate responses to resistance exercise and thereafter, muscular adaptation. Although the importance of the mechanical and hormonal stimulus to strength and power adaptation is well recognised, the contribution of the metabolic stimulus is largely unknown. Relatively few studies for example, have examined metabolic change across neuronal and dynamic power schemes, and not withstanding the fact that those mechanisms underpinning muscular adaptation, in relation to the metabolic stimulus, remain highly speculative. Inconsistent findings and methodological limitations within research (e.g. programme design, sampling period, number of samples) make interpretation further difficult. We contend that strength and power research needs to investigate those metabolic mechanisms likely to contribute to weight-training adaptation. Further research is also needed to examine the metabolic responses to different loading schemes, as well as interactions across age, sex and training status, so our understanding of how to optimise strength and power development is improved.

Biotransformation↗
PubMed1995

[Intracerebral microdialysis and its application in neurosciences].

As a new technique for sampling the extracellular fluid in vivo, the microdialysis is recently attractive to neuroscientists. Coupled with high sensitivity techniques for chemical microanalysis, it has been successfully used for monitoring chemical changes in extracellular environment of intact brain, and applied to some fields of neuroscience studies. In this article, its principle and some methodological aspects were described systematically, and a brief overview of its applications in neurosciences and some new advances was also given. Besides, the scope and limitations of microdialysis was discussed in the light of our own experience.

Animals↗
PubMed2003

Background and educational characteristics of prelingually deaf children implanted by five years of age.

PURPOSE: This study documents child, family and educational characteristics of a large representative sample of 8- to 9-yr-old prelingually deaf children who received a cochlear implant by 5 yr of age. Because pre-existing factors such as the child's gender, family characteristics, additional handicaps, age at onset of deafness and at implant, may affect postimplant outcomes, these variables must be accounted for before the impact of educational factors on performance with an implant can be adequately determined. Classroom variables that may affect postimplant outcomes include placement in public or private, mainstream or special education, oral or total communication environments. Other intervention variables include type and amount of individual therapy, experience of the therapist and parent participation in therapy. Documenting these characteristics for a large representative sample of implanted children can provide clinicians and researchers with insight regarding the types of families who sought early cochlear implantation for their children and the types of educational programs in which they placed their children after implantation. It is important to undertake studies that control for as many of these factors as possible so that the relative benefits of specific educational approaches for helping children to get the most benefit from their cochlear implant can be identified. METHOD: Over a 4-yr period, 181 children from across the US and Canada, accompanied by a parent, attended a cochlear implant research camp. Parents completed questionnaires in which they reported the child's medical and educational history, characteristics of the family, and their participation in the child's therapy. The parent listed names and addresses of clinicians who had provided individual speech/language therapy to the child and signed permission for these clinicians to complete questionnaires describing this therapy. RESULTS: To the extent that this sample is representative of those families seeking a cochlear implant for their child, especially during the initial period of device availability, this population can be characterized as follows. Most parents had normal hearing, were of majority (white) ethnicity and had more education and higher incomes than the general population. The families tended to be intact with both a mother and a father who involved their hearing-impaired child in family activities on a regular basis. The children were enrolled in the full range of educational placements available across the United States and Canada. Fairly even distributions of children from public and private schools, special education and mainstream classes and oral and total communication methodologies were represented. Educational placement changed as children gained increased experience with a cochlear implant. They received an increased emphasis on speech and auditory skills in their classroom settings and tended to move from private school and special education settings to public school and mainstream programs. These data support the position that early cochlear implantation is a cost effective procedure that allows deaf children to participate in a normal school environment with hearing age mates.

Canada↗
PubMed1995

Beliefs and perceptions of childbearing women choosing different primary health care providers.

The beliefs and perceptions of 60 low-risk childbearing women (n = 60) who had either nurse midwives or obstetricians as health care providers are identified in this descriptive phenomenological study. Interviews were conducted following childbirth, and participants completed the Utah Test for the Childbearing Year, which measures beliefs and perceptions about childbearing. There were no significant demographic differences, or significant differences in fear of childbirth, between groups of women having different providers. Mothers with physician caregivers had significantly higher rates of epidural anesthesia for pain management, higher levels of reliance on others, and less active participation in childbirth care decisions. Mothers with certified nurse midwives as caregivers participated more actively in childbirth care decisions and emphasized the quality of the birth experience. In this sample, differences in the beliefs and perceptions of women choosing different health care providers are identified. The importance of a philosophical fit with care providers and availability of options that are congruent with the individualized needs of childbearing women is evident from the study findings.

Adolescent↗
PubMed1982

Equilibrium dialysis for determination of protein binding or imipramine--evaluation of a method.

A range of methodological problems associated with equilibrium dialysis for serum or plasma protein binding assay of a weakly basic drug imipramine, in man has been carried out in order to establish a reliable procedure for large scale studies. Freezing of serum samples did not change the binding significantly, but a slight tendency towards increased binding to samples stored for 30 days was found. Protein binding was significantly higher in serum than in heparinized or citrated plasma, whereas there was no significant difference between plasma obtained by use of heparin or citrate as anticoagulant. There was a linear correlation between temperature (25-42 degrees) and binding such that binding decreased 0.1% per centigrade increase in temperature. Increase in pH resulted in increase in binding (0.6% per 0.1 unit pH). Equilibrium between serum and buffer phase was obtained within 5-6 hrs. The changes following osmotic equilibrium necessitated volume corrections of about 4-5% after 5-6 hrs, and 20-25% after 16-20 hrs (overnight experiments). With this correction the free fraction estimate remained constant for 26 hrs. Application of small volumes in the dialysis chamber may make the volume correction difficult and inaccurate. The use of Tris-Ringer buffer gave free fraction values about 20% higher (absolute values of free fraction) than that with phosphate buffer. With different imipramine concentrations a constant degree of binding was found at "therapeutic" levels, whereas a significantly decreased binding was found at a concentration of 1000 ng/ml. Control of these methodological factors is a prerequisite for obtaining reproducible and valid results, and authors should give more details on methodology in this field.

Blood Proteins↗
PubMed2006

Analysis of anatoxin-a in biological samples using liquid chromatography with fluorescence detection after solid phase extraction and solid phase microextraction.

Anatoxin-a is a naturally occurring, potent neurotoxin produced by some species of cyanobacteria in freshwaters. This toxin, which is a potential health hazard, especially to animals, has been determined in different biological matrices such as several cyanobacterial cultures and water samples and carps and mussels tissue using a sensitive High Performance Liquid Chromatography with Fluorescence detection method. Sonication was the technique selected for the extraction of intracellular anatoxin-a and solid phase extraction using weak cation exchange was used for the concentration and purification of the samples. 4-Fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F) was used to convert anatoxin into a highly fluorescent derivative. Recovery experiments were performed for each type of matrix used in this work, and adequate values were obtained (71-87%). Limits of detection for anatoxin-a were estimated to be in the ng/L and ng/g level for water and cyanobacterial samples, respectively. The results obtained were also compared with those obtained after using solid phase microextraction, as an alternative for the extraction and purification of the samples. Advantages and disadvantages regarding to the efficiency for impurities removal, simplicity and rapidity and the potential for concentration enhancement of using both methodologies have been also discussed.

Chromatography, High Pressure Liquid↗