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PubMed2005

Cell microarray for screening feeder cells for differentiation of embryonic stem cells.

Microarrays are currently recognized as one of major tools in the assessment of gene expression via cDNA or RNA analysis and are now accepted as a powerful experimental tool for high-throughput screening of a large number of samples, such as cDNA and siRNAs. In this study, we examined the potential of the microarray methodology for high-throughput screening of candidate cells as feeder cells which effectively differentiate embryonic stem (ES) cells to the specific lineage. Cell arrays were prepared by applying three kinds of cells, PA6, human umbilical vein endothelial, and COS-1 cells, to circular spots, 2 mm in diameter, on a glass plate, followed by the application of mouse ES cells to the cell microarray. After 8 d in culture, TuJ1 (neuron-specific class III beta-tubulin) immunocytochemical staining clearly demonstrated that only PA6 cell spots had the capability to induce ES cells to neuronal differentiation. Although this is a model experiment, these findings clearly indicate that the cell microarray will become a powerful tool for high-throughput screening large numbers of candidate feeder cells for specific differentiation.

Animals↗

PubMed2004

Modeling of the signals of an optical particle counter for real nonspherical particles.

An optical particle counter (OPC) was exposed to atmospheric particles of diameters of 200, 300, and 400 nm. The OPC data were combined with the results of single-particle analysis with a transmission electron microscope (TEM) on samples taken in parallel with the OPC measurements. With a T-matrix-based optical model the measured OPC spectra of scattered light pulses could be approximated with good precision. With an algorithm that simulated the response of the OPC to a given population of model particles derived from the TEM results, average absorption properties of different particle types were retrieved. For mobility sizes of 400 nm, higher light absorption was retrieved with the optical model for soot aggregates than for the rest of the morphological particle types. At smaller mobility sizes no compositional information could be derived from the model particles derived from the TEM data. Despite the limited success of the new methodology applied to the present experiment the results encourage the use of OPCs in combination with electrical mobility analyzers to derive more than aerosol-size distributions. With state-of-the-art pulse-height analysis the light-scattering pulses could be resolved with much finer resolution than in the instrument used.

Aerosols↗

PubMed2002

Patients' subjective experiences of antipsychotics: clinical relevance.

The subjective experience of patients with schizophrenia who are receiving antipsychotic medication has been a neglected research area, as has the satisfaction of patients with their drug treatments. This is unfortunate, as satisfaction with treatment appears to be related strongly to the readiness of patients to take their medication as prescribed, and thereby to outcome. Patients' perceptions of their treatment do not appear to be related strongly to severity of illness or symptom ratings, although there are associations between perceptions of treatment and adverse effects. Surveys of patient experience with typical antipsychotics have tended to indicate high levels of dissatisfaction and perceived adverse effects. There have been a number of surveys of patients' perceptions of treatment with the atypical antipsychotics. These tend to accord with the expectation that a relative freedom from adverse effects with the atypical antipsychotics will be reflected in enhanced levels of satisfaction and perceived well-being. In general, these studies share a number of weaknesses, including small sample sizes, bias in selection of respondents, open treatment and lack of suitable comparator groups. In addition, many have adopted a cross-sectional, rather than longitudinal, approach and have relied on nonvalidated and perhaps idiosyncratic rating measures. Recently, there have been studies of better methodological quality. These, too, have indicated that patients regard the newer treatments more positively than the older regimens. In addition, there is now evidence that the various new-generation antipsychotics may be evaluated differently by patients.

Antipsychotic Agents↗

PubMed2003

External quality assurance program for PCR amplification of genomic DNA: an Italian experience.

BACKGROUND: External quality assurance (EQA) programs for diagnostic tests based on nucleic acid amplification have not been widely implemented in clinical laboratories and remain limited to few tests. Development of specific EQA programs based on application-based proficiency testing for any diagnostic molecular target is challenging. Development of EQA trials based on methodologic proficiency testing and directed to the evaluation of analytical aspects common to the majority of PCR-based tests may be valuable. METHODS: We developed an EQA program for evaluation of DNA extraction and amplification and analysis of products after PCR. Participants received a package containing primers and reference materials to evaluate three specific controls for, respectively, DNA extraction (quality and quantity), PCR performance (specificity and efficiency), and interpretation of results after electrophoresis. Each participant was asked to return to the organizers a form with their numerical results and an aliquot of all amplified samples for joint evaluation. RESULTS: Results varied in all phases of the experimental procedure: preamplification, amplification, and post-PCR interpretation. To give a general estimation on the quality of performances for each laboratory, we designed a score scheme in which the results of any specific action were evaluated on the basis of the distribution around the median consensus values. The maximum possible score was 84. On the basis of total score obtained by each laboratory, we created a qualitative ranking list that provided the final interpretation of results as excellent (>63 points; n = 4 laboratories), good (53-63 points; n = 13), sufficient (42-52 points; n = 15), poor (31-41 points; n = 3), and not acceptable (<31 points; n = 4). CONCLUSIONS: This survey demonstrates the importance of EQA trials based on methodologic proficiency testing directed to evaluation of analytical aspects common to the majority of PCR-based tests.

DNA↗

PubMed1993

Growth versus stability: older primiparous women as a paradigmatic case for persistence.

The early pregnancy experience of women over 30 offers paradigmatic evidence which supports the significance of persistence for the study of nursing phenomenon. Pregnancy and first-time motherhood have come to be associated with change, growth and development. This research, which was undertaken to describe the changes experienced by older primiparous women during early pregnancy, illustrates how research grounded in a particular paradigm, in this case the growth or developmental paradigm, can limit the interpretation of data. When viewed from a developmental perspective, women in this sample were seen as resistant to change and could be said to be having trouble 'adapting' to their impending motherhood. However, when these women were viewed from the perspective of stability they were seen to engage in activities that allowed them to persist in their current self-view. In the biological sciences, persistence refers to a compensatory process which allows a system to maintain goal-directedness in spite of both internal and external disturbances. Women in this sample used balancing and buffering to continue their self-patterns. Balancing represented the verbal and behavioural processes women engaged in to integrate their experience of themselves as pregnant into their sense of themselves as not pregnant. Buffering included those activities that women used to maintain a sense of themselves in the face of the threat posed by their impending motherhood. Balancing and buffering represent mechanisms used in the service of persistence and reflect pattern maintenance activities.

Adaptation, Psychological↗

PubMedDate not supplied

The lived experience of adult male survivors who allege childhood sexual abuse by clergy.

This phenomenological study describes the essential structure of the lived experience of adult male survivors of childhood sexual abuse by clergy (AMSCSABC). A purposive sample of seven AMSCSABC related their subjective experiences in semistructured interviews. Colaizzi's (1978) phenomenological method was used for data analysis. Survivors describe a bifurcated rage and spiritual distress that pervades their entire "lifebeing." Learning about AMSCSABC will assist nurses to identify potential risk factors of childhood sexual abuse by clergy (CSABC), design prevention strategies, and enhance empathy for a healing relationship.

Adaptation, Psychological↗

PubMed2002

Experience of the first survey of the pilot project of Asian Quality Assurance Survey program.

The Asian Network for Clinical Laboratory Standardization (ANCLS) decided to start her First Inter-laboratory Quality Assurance survey at the business meeting during the Second Asian Colloquium on October 21-22, 2000 in Kobe, Japan. The first survey materials of Asian Quality Assurance Survey (AQuAS) were distributed in July 2001 to 21 hospitals in the field of chemistry and 22 hospitals in the field of hematology among total 24 participating laboratories from seven Asian countries: Indonesia, Japan, Korea, Malaysia, Philippines, Singapore and Thailand. The survey methods in chemistry and in hematology were somewhat different. In chemistry the survey material was considered as unknown but handled similarly to the sample from the patient. Nineteen routine tests were performed only once. The hematology samples were considered as unknown but handled similarly to control or calibration material. Five parameters were tested five times repetitively and their average for each parameter was calculated. All the results were supposed to be sent back to the office within two weeks. Although it took more than two weeks, the return rate was 100%. The analysis was performed in several ways such as all the results together, by instruments and by methods. Mean, standard deviation (SD), standard deviation index (SDI), coefficient of variation (CV) and variance index score (VIS) were to be calculated in chemistry, and in hematology the same parameter were to be calculated except CV and VIS. In the first survey, the CV in chemistry was not calculated and the analysis by instrument or by methodology was also not attempted since there were not enough participating hospitals to do such analysis. In hematology the analysis was done by instrument only. The survey process was carried out successfully though there were some difficulties in communication tools, transportation methods and handling of specimens due to different weather conditions, and returning the report in the correct unit and to the correct place. The submitted data were acceptable for analysis. There were some differences in the units of measurement in different countries or laboratories. It was necessary to convert some of the units. Some laboratories apparently do not perform certain tests such as calcium, potasium and gamma-glutamyltransferase (gamma-GT). The gamma-GT is the most frequently not performed test. With the experience of this first survey, all the members involved in the survey have been trained well to do future surveys.

Asia↗

PubMed2004

A complete set of novel 2D correlation NMR experiments based on heteronuclear J-cross polarization.

A suite of multiple-purpose sensitivity-enhanced 2D correlation NMR experiments based on heteronuclear J-cross polarization (HCP) techniques are introduced for isotropic liquid samples. Several pulse sequences using an adaptable heteronuclear TOCSY mixing building block are proposed for different types of effective coherence-order-selective (COS) heteronuclear coherence-transfer mechanisms. They are based on the anisotropic behaviour of the involved HCP process that is easily described and analysed in terms of cartesian product-operator formalism. A number of different versions are given for in-phase to in-phase (II-COS: S (-) \\ to I (-)), in-phase to anti-phase (IA-COS: S (-) \\ to 2 I (-) S (z)), in-phase to spin-state-selective (IS(3)-COS: S (-) \\ to 2 I (-) S (alpha /beta)), anti-phase to in-phase (AI-COS: 2 I (z) S (-) \\ to I (-)), anti-phase to anti-phase (AA-COS: 2 I (z) S (-) \\ to 2 I (-) S (z)), anti-phase to spin-state-selective (AS(3)-COS: 2 I (z) S (-) \\ to 2 I (-) S (alpha /beta)) and spin-state-selective to spin-state-selective (S(3)S(3)-COS: 2 I (alpha /beta) S (-) \\ to 2 I (-) S (alpha /beta )) coherence transfers. The combination of the echo/anti-echo approach, heteronuclear gradient echoes and the preservation of equivalent pathways (PEP) methodology affords a general approach to obtain sensitivity-enhanced pure-absorption 2D spectra that can be used as interesting alternatives to conventional pulse-interrupted free-precession INEPT-based pulse schemes, such as HSQC-type and TROSY-type experiments.

Anisotropy↗

PubMed1996

The challenge project: perpetrators of child sexual abuse in south east London.

The collection of descriptive data on sex offenders has been marred by methodological problems: failing to distinguish between different types of sex offenders, or using selective samples form specialist institutions. The Challenge Project is a community project for perpetrators of child sexual abuse in South East London. Over a 2 year period, all convicted child molesters appearing before the court for sentencing or applying for parole were assessed by the Project Team. Data was collected on a range of personal, offense-related, and psychometric variables. Some of these were replicated on violent and property offender populations to allow for a preliminary comparison to be made. The descriptive profile of the offenders mirrored findings of previous research, and the comparison with other offender groups suggested that there is a cluster of childhood experiences which differentiate these sex offenders from other types of offenders. The research findings supported the hypothesis that perpetrators of child sexual abuse from a heterogeneous group of offenders, with identifiable--albeit overlapping--subgroups related to key variables such as a history of sexual abuse, repeat offending, and victim gender and relationship. The essential importance of these variables in relation to compliance with and response to treatment is discussed in further research.

Adolescent↗

PubMed1998

Coping in young women: theoretical retroduction.

Although there is increasing interest in women's health, there remains little empirical evidence concerning concepts related to health and women's life trajectory. Our purpose in conducting this research was to examine women's developmental stressors and coping during young adulthood. Lazarus and Folkman's coping model provided sensitizing concepts for data generation in a retroductive research methodology. The sample included 26 women of diverse perspectives who participated in interviews and groups. Philosophical concerns of subjectivity, context and gender sensitivity were actualized in iterative stages of data collection, analysis and comparison to extant theory. Rapidly expanding multiple roles were identified as developmental stressors for young women. Decontextualized coping strategies are arranged on a behavioural continuum: avoidance, distraction, and confrontation. Contextualized descriptions of coping are presented in two representative accounts: Lauren and Mary Ellen. A philosophical stance, assumed at about the age of 30, provides a lens for assessing developmental stressors that influence the appraisal process. Rich descriptions of young women's coping and developmental influences move beyond abstract theory to a contextual understanding of young women's experiences.

Adaptation, Psychological↗

PubMed2004

Microdialysis of the brain structures: application in behavioral research on vasopressin and oxytocin.

In vivo microdialysis allows sampling of brain regions in conscious, freely moving animals. Moreover, the in vivo microdialysis allows to administer drugs directly into specific brain areas. Both are useful in behavioral studies. The subject of this review is the methodology of brain microdialysis, that is construction of the probe, effect of temperature, composition of the perfusion medium, perfusion flow rate, characteristics of the membrane material and the role of the diffusion coefficient. Other techniques of the study of in vivo release, alternative for microdialysis, are described. Advantages and disadvantages of acute and chronic microdialysis are discussed. Chronic microdialysis is especially needed in behavioral studies. Finally, the examples of application of microdialysis in behavioral studies of vasopressin (AVP) and oxytocin (OXT) and our own experience in these studies are described.

Animals↗

PubMed2003

An overview of qualimetric strategies for optimisation and calibration in pharmaceutical analysis using flow injection techniques.

Flow Injection analysis represents an attractive tool because of its great advantages, such as versatility, speed, high sampling rate and wide applicability in the field of pharmaceutical analysis. However, due to the inherent characteristics of the technique, the choice of the best set of operational and chemical conditions is complicated and the conventional univariate optimisation method present some limitations, mainly due to fact that the interdependence of variables is not considered. In relation to the calibration process, because of the transient character of the signals obtained using FIA manifolds coupled with different detection techniques, different strategies can be used in calibration to solve some problems related to the nature of the signal thereby improving the performance characteristics of the method. This paper offers an overview of different methodologies used in optimisation based on the use of statistically designed experiments and some strategies developed for calibration applied to the analysis of pharmaceuticals.

Calibration↗

PubMed1991

Precise gene dosage determination by polymerase chain reaction: theory, methodology, and statistical approach.

We developed a general method of quantifying relative copy numbers of specific DNA sequences based on the theoretical accumulation of polymerase chain reaction (PCR) products when two DNA sequences are amplified together (co-amplified). Our experiments illustrate the development and theory of the technique. The precision of our estimates is demonstrated by statistical confidence intervals. Tests for effects introduced by experimental factors were performed. The precision of the technique was established by examining the relative gene dosage of the X-linked dystrophin gene in human genomic DNAs from a male, a normal female, a 47,XXX female, and a 48,XXXX cell line. The sensitivity was sufficient to distinguish three copies of the gene from four copies; equivalent to detecting loss of heterozygosity in half the cells of a tumour. Confidence intervals allowed us to reject the hypothesis that there was no difference between DNA samples. Four sample pairs would be required to demonstrate relative gene dosage ratios of 2.0 to 1.0; eight sample pairs would be required to demonstrate a relative gene dosage ratio of 1.3 to 1.0. This method should be useful in detecting gene amplification and deletion in a variety of situations.

Base Sequence↗

PubMed2000

A non-radioisotopic quantitative competitive polymerase chain reaction method: application in measurement of human herpesvirus 7 load.

Quantitative-competitive polymerase chain reaction (QCPCR) is a well-optimised and objective methodology for the determination of viral load in clinical specimens. A major advantage of QCPCR is the ability to control for the differential modulation of the PCR process in the presence of potentially inhibitory material. QCPCR protocols were developed previously for CMV, HHV-6, HHV-7 and HHV-8 and relied upon radioactively labelled primers, followed by autoradiography of the separated and digested PCR products to quantify viral load. Whilst this approach offers high accuracy and dynamic range, non-radioactive approaches would be attractive. Here, an alternative detection system is reported, based on simple ethidium bromide staining and computer analysis of the separated reaction products, which enables its adoption in the analysis of a large number of samples. In calibration experiments using cloned HHV-7 DNA, the ethidium bromide detection method showed an improved correlation with known copy number over that obtained with the isotopic method. In addition, 67 HHV-7 PCR positive blood samples, derived from immunocompromised patients, were quantified using both detection techniques. The results showed a highly significant correlation with no significant difference between the two methods. The applicability of the computerised densitometry method in the routine laboratory is discussed.

DNA, Viral↗

PubMed2003

Determination of Pb complexation in oxic and sulfidic waters using pseudovoltammetry.

Pseudovoltammetry was used to evaluate the actual Pb complexation occurring in natural water samples of varying oxygen and sulfide concentration. In pseudovoltammetry, the potential at which metal-ligand complexes are broken up to form the metal amalgam is used to determine the complexes' thermodynamic stability constants (KTHERM; corrected for metal and ligand side reaction coefficients) via the Nernst expression. This methodology removes the need for any metal additions and for subsequent modeling using fitting criteria, which provide only conditional stability constant data (KCOND). Using known organic ligands, a chelate scale ranging from log KTHERM = 4 to log KTHERM = 20 was developed as a template for comparison with samples collected from two stations of different salinities and at several depths in the Chesapeake Bay. These samples were observed to contain up to five different ligand compounds of unknown structural composition (log KTHERM > 8) with the strongest ligand fraction exceeding log KTHERM > 39 (the maximum observable thermodynamic stability constant due to the reduction of Na+). One possible explanation for the observed complexation is the existence of lead sulfide clusters. This was supported by laboratory experiments using electrochemistry and ICR-FTMS, which confirmed the formation of electrochemically inert multinuclear clusters with high stability constants (e.g., M3S3, log KTHERM = 62.9). However, in all field samples, (sub)nanomolar levels of acid-leachable sulfide were recovered at pH 5.0-6.2, which could be attributed to dissociation of lead sulfide complexes with moderate acidity. Recovery of sulfide increased from < 10% of the total dissolved Pb concentration (Pbdiss) in surface waters to 100% of the Pbdiss in the sulfide-rich bottom waters at the higher salinity location.

Electrochemistry↗

PubMed2004

Determination of sugar structures in solution from residual dipolar coupling constants: methodology and application to methyl beta-D-xylopyranoside.

We have developed methodology for the determination of solution structures of small molecules from residual dipolar coupling constants measured in dilute liquid crystals. The power of the new technique is demonstrated by the determination of the structure of methyl beta-d-xylopyranoside (I) in solution. An oriented sample of I was prepared using a mixture of C(12)E(5) and hexanol in D(2)O. Thirty residual dipolar coupling constants, ranging from -6.44 to 4.99 Hz, were measured using intensity-based J-modulated NMR techniques. These include 15 D(HH), 4 (1)D(CH), and 11 (n)D(CH) coupling constants. The accuracy of the dipolar coupling constants is estimated to be < +/- 0.02 Hz. New constant-time HMBC NMR experiments were developed for the measurement of (n)D(CH) coupling constants, the use of which was crucial for the successful structure determination of I, as they allowed us to increase the number of fitted parameters. The structure of I was refined using a model in which the directly bonded interatom distances were fixed at their ab initio values, while 16 geometrical and 5 order parameters were optimized. These included 2 CCC and 6 CCH angles, and 2 CCCC and 6 CCCH dihedral angles. Vibrationally averaged dipolar coupling constants were used during the refinement. The refined solution structure of I is very similar to that obtained by ab initio calculations, with 11 bond and dihedral angles differing by 0.8 degrees or less and the remaining 5 parameters differing by up to 3.3 degrees . Comparison with the neutron diffraction structure showed larger differences attributable to crystal packing effects. Reducing the degree of order by using dilute liquid crystalline media in combination with precise measurement of small residual dipolar coupling constants, as shown here, is a way of overcoming the limitation of strongly orienting liquid crystals associated with the complexity of (1)H NMR spectra for molecules with more than 12 protons.

Carbohydrate Conformation↗

PubMed2001

Assessing culture via the Internet: methods and techniques for psychological research.

This study examines the acculturation experiences of Arabic immigrants and assesses the utility of the Internet as a data collection tool. Based on in-depth pilot interview data from 10 male Arabic immigrants and items selected from pre-existing measures, the Male Arabic Ethnic Identity Measure (MAEIM) was developed. Male Arab immigrants (115 males) were solicited through traditional methods in addition to the Internet. Satisfactory reliability and validity were reported for the MAEIM. No significant differences emerged between the Internet and Midwestern samples. The Internet proved to be an effective method for soliciting a relatively large, geographically dispersed sample of Arabic immigrants. The use of the Internet as a research tool is examined in the context of anonymity, networking, low-cost, perceived interactive control, methodological rigor, and external validity. The Internet was an effective vehicle for addressing concerns raised by prospective participants. It is suggested that the Internet may be an important method to assess culture-relevant variables in further research on Arab and other immigrant populations.

Acculturation↗

PubMed2005

Predictive model for reduction of Escherichia coli during acetic acid decontamination of chicken skin.

AIMS: The response surface methodology was used to evaluate the effect of operating variables (acetic acid concentration, spraying time and temperature) on the reduction of Escherichia coli populations on poultry breast skin in a laboratory showering process, as well as to identify the best conditions that are required to develop this operation. METHODS AND RESULTS: Skin samples were inoculated with a 24-h E. coli culture and afterwards treated according to experimental design under selected acetic acid concentration, spraying time, and solution temperature. The E. coli reduction model was significantly affected by the acetic acid concentration and spraying time (P < or = 0.05 and < or =0.01), while temperature did not show a significant effect (P > 0.05). CONCLUSION: The predictive model obtained was validated through additional confirmatory experiments and showed to be adequate, and it could be used as an approach to optimize the acetic acid spray washes during poultry carcasses processing. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of acetic acid washes in the processing of poultry does not have the capability of eliminating E. coli populations from carcasses. However, significant reductions in the initial load could be achieved.

Acetic Acid↗