Use of accuracy profile for the validation of a gas chromatography-negative chemical ionization tandem mass spectrometry method: quantification of ethyl glucuronide in hair

Introduction: Ethylglucuronide (EtG) is a direct and specific metabolite of ethanol. Its determination in hair is of increasing interest for detecting and monitoring alcohol abuse. The quantification of EtG in hair requires analytical methods showing highest sensitivity and specificity. We present a fully validated method based on gas chromatography-negative chemical ionization tandem mass spectrometry (GC-NCI-MS/MS). The method was validated using French Society of Pharmaceutical Sciences and Techniques (SFSTP) guidelines which are based on the determination of the total measurement error and accuracy profiles. Methods: Washed and powdered hair is extracted in water using an ultrasonic incubation. After purification by Oasis MAX solid phase extraction, the derivatized EtG is detected and quantified by GC-NCI-MS/MS method in the selected reaction monitoring mode. The transitions m/z 347 / 163 and m/z 347 / 119 were used for the quantification and identification of EtG. Four quality controls (QC) prepared with hair samples taken post mortem from 2 subjects with a known history of alcoholism were used. A proficiency test with 7 participating laboratories was first run to validate the EtG concentration of each QC sample. Considering the results of this test, these samples were then used as internal controls for validation of the method. Results: The mean EtG concentrations measured in the 4 QC were 259.4, 130.4, 40.8, and 8.4 pg/mg hair. Method validation has shown linearity between 8.4 and 259.4 pg/mg hair (r2 > 0.999). The lower limit of quantification was set up at 8.4 pg/mg. Repeatability and intermediate precision were found less than 13.2% for all concentrations tested. Conclusion: The method proved to be suitable for routine analysis of EtG in hair. GC-NCI-MS/MS method was then successfully applied to the analysis of EtG in hair samples collected from different alcohol consumers.

Validation of a modified fluorimetric assay for the screening of trichomonacidal drugs

A fluorimetric microassay that uses a redox dye to determine the viability of the flagellate Trichomonas vaginalis has been optimised to provide a more sensitive method to evaluate potential trichomonacidal compounds. Resazurin has been used in recent years to test drugs against different parasites, including trichomonadid protozoa; however, the reproducibility of these resazurin-based methods in our laboratory has been limited because the flagellate culture medium spontaneously reduces the resazurin. The objective of this work was to refine the fluorimetric microassay method previously developed by other research groups to reduce the fluorescence background generated by the media and increase the sensitivity of the screening assay. The experimental conditions, time of incubation, resazurin concentration and media used in the microtitre plates were adjusted. Different drug sensitivity studies against T. vaginalis were developed using the 5-nitroimidazole reference drugs, new 5-nitroindazolinones and 5-nitroindazole synthetic derivatives. Haemocytometer count results were compared with the resazurin assay using a 10% solution of 3 mM resazurin dissolved in phosphate buffered saline with glucose (1 mg/mL). The fluorimetric assay and the haemocytometer counts resulted in similar percentages of trichomonacidal activity in all the experiments, demonstrating that the fluorimetric microtitre assay has the necessary accuracy for high-throughput screening of new drugs against T. vaginalis.

Validation transculturelle de l'Oswestry disability index en français [Cross-cultural validation of the Oswestry disability index in French]

AIM: The aim of this study was to interpret and validate a French version of the Oswestry disability index (ODI), using a cross-cultural validation method. The validity and reliability of the questionnaire was assessed in order to ensure the psychometric characteristics. METHOD: The cross-cultural validation was carried out according to Beaton's methodology. The study was conducted with 41 patients suffering from low back pain. The correlation between the ODI and the Roland-Morris disability questionnaire (RMDQ), the medical outcome survey short form-36 (MOS SF-36) and a pain visual analogical scale (VAS) was assessed. RESULTS: The validity of the Oswestry questionnaire was studied using the Cronbach Alpha coefficient calculation: 0.87 (n=36). The significant correlation between the ODI and RMDQ was 0.8 (P<0.001, n=41) and 0.71 (P<0.001, n=36) for the pain VAS. The correlation between the ODI and certain subscales (physical functioning 0.7 (P<0.001, n=41), physical role 0.49 et bodily pain 0.73 (P<0.001, n=41)) of the MOS SF-36 were equally significant. The reproducibility of the ODI was calculated using the Wilcoxon matched pairs test: there was no significant difference for eight out of ten sections or for the final score. CONCLUSION: This French translation of the ODI should be considered as valid and reliable. It should be used for any future clinical studies carried out using French language patients. Complimentary studies must be completed in order to assess its sensitivity to change in the event of any modifications in the patients functional capacity.

Quality of life after pulmonary embolism: validation of the French version of the PEmb-QoL questionnaire.

Background The PEmb-QoL is a validated 40-item questionnaire to quantify health-related quality of life in patients having experienced pulmonary embolism (PE). It covers six health dimensions: frequency of complaints, activities of daily living limitations, work-related problems, social limitations, intensity of complaints, and emotional complaints. Originally developed in Dutch and English, we sought to prospectively validate the psychometric properties of a French version of the PEmb-QoL.MethodsWe performed a forward and backward translation of the English version of the PEmb-QoL into French. French-speaking consecutive adult patients with an acute, objectively confirmed PE admitted to the emergency department of a Swiss university hospital between 08/2009 and 09/2011 were recruited telephonically. We used standard psychometric tests and criteria to evaluate the acceptability, reliability, and validity of the French version of the PEmb-QoL. We also performed an exploratory factor analysis.ResultsOverall, 102 patients were enrolled in the study. The French version of the PEmb-QoL showed good reliability (internal consistency, item¿total and inter-item correlations), reproducibility (test-retest reliability), and validity (convergent, discriminant) in French-speaking patients with PE. The exploratory factor analysis suggested three underlying dimensions: limitations in daily activity (items 4b-m, 5a-d), symptoms (items 1a-h and 7), and emotional complaints (items 9a-f and j).ConclusionWe successfully validated the French version of the PEmb-QoL questionnaire in patients with PE. Our results show that the PEmb-QoL is a valuable tool for assessing health-related quality of life after PE in French-speaking patients.

Development and validation of AMANDA, a new algorithm for selecting highly relevant regions in Molecular Interaction Fields

Descriptors based on Molecular Interaction Fields (MIF) are highly suitable for drug discovery, but their size (thousands of variables) often limits their application in practice. Here we describe a simple and fast computational method that extracts from a MIF a handful of highly informative points (hot spots) which summarize the most relevant information. The method was specifically developed for drug discovery, is fast, and does not require human supervision, being suitable for its application on very large series of compounds. The quality of the results has been tested by running the method on the ligand structure of a large number of ligand-receptor complexes and then comparing the position of the selected hot spots with actual atoms of the receptor. As an additional test, the hot spots obtained with the novel method were used to obtain GRIND-like molecular descriptors which were compared with the original GRIND. In both cases the results show that the novel method is highly suitable for describing ligand-receptor interactions and compares favorably with other state-of-the-art methods.

Validation of the Addiction Severity Index in French-speaking alcoholic patients.

OBJECTIVE: Evaluation of a French translation of the Addiction Severity Index (ASI) in 100 (78 male) alcoholic patients. METHOD: Validity of the instrument was assessed by measuring test-retest and interrater reliability, internal consistency and convergence and discrimination between items and scales. Concurrent validity was assessed by comparing the scores from the ASI with those obtained from three other clinimetric instruments. RESULTS: Test-retest reliability of ASI scores (after a 10-day interval) was good (r = 0.63 to r = 0.95). Interrater reliability was evaluated using six video recordings of patient interviews. Severity ratings assigned by six rates were significantly different (p < .05), but 72% of the ratings assigned by those who viewed the videos were within two points of the interviewer's severity ratings. Cronbach alpha coefficient of internal consistency varied from 0.58 to 0.81 across scales. The average item-to-scale convergent validity (r value) was 0.49 (range 0.0 to 0.84) for composite scores and 0.35 (range 0.00 to 0.68) for severity ratings, whereas discriminant validity was 0.11 on average (range-0.19 to 0.46) for composite scores and 0.12 (range-0.20 to 0.52) for severity ratings. Finally, concurrent validity with the following instruments was assessed: Severity of Alcoholism Dependence Questionnaire (40% shared variance with ASI alcohol scale), Michigan Alcoholism Screening Test (2% shared variance with ASI alcohol scale) and Hamilton Depression Rating Scale (31% shared variance with ASI psychiatric scale). CONCLUSIONS: The Addiction Severity Index covers a large scope of problems encountered among alcoholics and quantifies need for treatment. This French version presents acceptable criteria of reliability and validity.

Clinical Learning Environment and Supervision. Development and Validation of the CLES Evaluation Scale

The purpose of the study is: (1) to describe how nursing students' experienced their clinical learning environment and the supervision given by staff nurses working in hospital settings; and (2) to develop and test an evaluation scale of Clinical Learning Environment and Supervision (CLES). The study has been carried out in different phases. The pilot study (n=163) explored the association between the characteristics of a ward and its evaluation as a learning environment by students. The second version of research instrument (which was developed by the results of this pilot study) were tested by an expert panel (n=9 nurse teachers) and test-retest group formed by student nurses (n=38). After this evaluative phase, the CLES was formed as the basic research instrument for this study and it was tested with the Finnish main sample (n=416). In this phase, a concurrent validity instrument (Dunn & Burnett 1995) was used to confirm the validation process of CLES. The international comparative study was made by comparing the Finnish main sample with a British sample (n=142). The international comparative study was necessary for two reasons. In the instrument developing process, there is a need to test the new instrument in some other nursing culture. Other reason for comparative international study is the reflecting the impact of open employment markets in the European Union (EU) on the need to evaluate and to integrate EU health care educational systems. The results showed that the individualised supervision system is the most used supervision model and the supervisory relationship with personal mentor is the most meaningful single element of supervision evaluated by nursing students. The ward atmosphere and the management style of ward manager are the most important environmental factors of the clinical ward. The study integrates two theoretical elements - learning environment and supervision - in developing a preliminary theoretical model. The comparative international study showed that, Finnish students were more satisfied and evaluated their clinical placements and supervision with higher scores than students in the United Kingdom (UK). The difference between groups was statistical highly significant (p= 0.000). In the UK, clinical placements were longer but students met their nurse teachers less frequently than students in Finland. Arrangements for supervision were similar. This research process has produced the evaluation scale (CLES), which can be used in research and quality assessments of clinical learning environment and supervision in Finland and in the UK. CLES consists of 27 items and it is sub-divided into five sub-dimensions. Cronbach's alpha coefficient varied from high 0.94 to marginal 0.73. CLES is a compact evaluation scale and user-friendliness makes it suitable for continuing evaluation.

Content validation of the dimensions constituting non-adherence to treatment of arterial hypertension

The objective of the study was to validate the content of the dimensions that constituted nonadherence to treatment of arterial systemic hypertension. It was a methodological study of content validation. Initially an integrative review was conducted that demonstrated four dimensions of nonadherence: person, disease/treatment, health service, and environment. Definitions of these dimensions were evaluated by 17 professionals, who were specialists in the area, including: nurses, pharmacists and physicians. The Content Validity Index was calculated for each dimension (IVCi) and the set of the dimensions (IVCt), and the binomial test was conducted. The results permitted the validation of the dimensions with an IVCt of 0.88, demonstrating reasonable systematic comprehension of the phenomena of nonadherence.

Development and validation of a clinical decision rule for the diagnosis of influenza.

INTRODUCTION: A clinical decision rule to improve the accuracy of a diagnosis of influenza could help clinicians avoid unnecessary use of diagnostic tests and treatments. Our objective was to develop and validate a simple clinical decision rule for diagnosis of influenza. METHODS: We combined data from 2 studies of influenza diagnosis in adult outpatients with suspected influenza: one set in California and one in Switzerland. Patients in both studies underwent a structured history and physical examination and had a reference standard test for influenza (polymerase chain reaction or culture). We randomly divided the dataset into derivation and validation groups and then evaluated simple heuristics and decision rules from previous studies and 3 rules based on our own multivariate analysis. Cutpoints for stratification of risk groups in each model were determined using the derivation group before evaluating them in the validation group. For each decision rule, the positive predictive value and likelihood ratio for influenza in low-, moderate-, and high-risk groups, and the percentage of patients allocated to each risk group, were reported. RESULTS: The simple heuristics (fever and cough; fever, cough, and acute onset) were helpful when positive but not when negative. The most useful and accurate clinical rule assigned 2 points for fever plus cough, 2 points for myalgias, and 1 point each for duration <48 hours and chills or sweats. The risk of influenza was 8% for 0 to 2 points, 30% for 3 points, and 59% for 4 to 6 points; the rule performed similarly in derivation and validation groups. Approximately two-thirds of patients fell into the low- or high-risk group and would not require further diagnostic testing. CONCLUSION: A simple, valid clinical rule can be used to guide point-of-care testing and empiric therapy for patients with suspected influenza.

The prognostic value of semi-quantitative i-123 mibg scintigraphy at diagnosis in high risk neuroblastoma: validation of the siopen score method

Purpose: SIOPEN scoring of 123I mIBG imaging has been shown to predict response to induction chemotherapy and outcome at diagnosis in children with HRN.Method: Patterns of skeletal 123I mIBG uptake were assigned numerical scores (Mscore) ranging from 0 (no metastasis) to 72 (diffuse metastases) within 12 body areas as described previously. 271 anonymised, paired image data sets acquired at diagnosis and on completion of Rapid COJEC induction chemotherapy were reviewed, constituting a representative sample of 1602 children treated prospectively within the HR-NBL1/SIOPEN trial. Pre-and post-treatment Mscores were compared with bone marrow cytology (BM) and 3 year event free survival (EFS).Results: Results 224/271 patients showed skeletal MIBG-uptake at diagnosis and were evaluable forMIBG-response. Complete response (CR) on MIBG to Rapid COJEC induction was achieved by 66%, 34% and 15% of patients who had pre-treatment Mscores of <18 (n¼65, 29%), 18-44 (n¼95,42%) and Y ´ 45 (n¼64, 28.5%) respectively (chi squared test p<.0001). Mscore at diagnosis and on completion of Rapid COJEC correlated strongly with BM involvement (p<0.0001). The correlation of pre score with post scores and response was highly significant (p<0.001). Most importantly, the 3 year EFS in 47 children with Mscore 0 at diagnosis was 0.68 (A ` 0.07), by comparison with 0.42 (A` 0.06), 0.35 (A` 0.05) and 0.25 (A` 0.06) for patients in pre-treatment score groups <18, 18-44 and Y ´ 45, respectively (p<0.001). AnMscore threshold ofY ´ 45 at diagnosis was associated with significantly worse outcome by comparison with all other Mscore groups (p¼0.029). The 3 year EFS of 0.53 (A` 0.07) of patients in metastatic CR (mIBG and BM) after Rapid Cojec (33%) is clearly superior to patients not achieving metastatic CR (0.24 (A ` 0.04), p¼0.005).Conclusion: SIOPEN scoring of 123I mIBG imaging has been shown to predict response to induction chemotherapy and outcome at diagnosis in children with HRN.

Validation of rotational thromboelastometry during cardiopulmonary bypass: A prospective, observational in-vivo study.

BACKGROUND: Rotational thromboelastometry (ROTEM) is a whole blood point-of-test used to assess the patient's coagulation status. Three of the available ROTEM tests are EXTEM, INTEM and HEPTEM. In the latter, heparinase added to the INTEM reagent inactivates heparin to reveal residual heparin effect. Performing ROTEM analysis during cardiopulmonary bypass (CPB) might allow the anaesthesiologist to anticipate the need for blood products. OBJECTIVE: The goal of this study was to validate ROTEM analysis in the presence of very high heparin concentrations during CPB. DESIGN: Prospective, observational trial. SETTING: Single University Hospital. PARTICIPANTS: Twenty patients undergoing coronary artery bypass grafting. MAIN OUTCOME MEASURE: ROTEM analysis was performed before heparin administration (T0), 10 min after heparin (T1), at the end of CPB (T2) and 10 min after protamine (T3). The following tests were performed: EXTEM, INTEM, and HEPTEM. Heparin concentrations were measured at T1 and at the end of bypass (T2). RESULTS: At T1, EXTEM differed from baseline for coagulation time: +26.7 s (18.4 to 34.9, P < 0.0001), α: -3° (1.0 to 5.4, P = 0.006) and A10: -4.4 mm (2.3 to 6.5, P = 0.0004). INTEM at T0 was different from HEPTEM at T1 for coagulation time: + 47 s (34.3 to 59.6, P >0.0001), A10: -2.3 mm (0.5 to 4.0, P = 0.01) and α -2° (1.0 to 3.0; P = 0.0007). At T2, all parameters in EXTEM and HEPTEM related to fibrin-platelet interaction deteriorated significantly compared to T1. At T3, EXTEM and INTEM were comparable to EXTEM and HEPTEM at T2. CONCLUSION: HEPTEM and EXTEM measurements are valid in the presence of very high heparin concentrations and can be performed before protamine administration in patients undergoing cardiac surgery with CPB. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT01455454.

Task-shifting in the delivery of hormonal contraceptive methods: Validation of a questionnaire and preliminary results.

Guilbert ER, Morin D, Guilbert AC, Gagnon H, Robitaille J, Richardson M. International Journal of Nursing Practice 2011; 17: 315-321 Task-shifting in the delivery of hormonal contraceptive methods: Validation of a questionnaire and preliminary results In order to palliate the access problem to effective contraceptive methods in Quebec, Canada, as well as to legitimate nurses' practices in family planning, a collaborative agreement was developed that allow nurses, in conjunction with pharmacists, to give hormonal contraceptives to healthy women of reproductive age for a 6 month period. Training in hormonal contraception was offered to targeted nurses before they could begin this practice. A questionnaire, based on Rogers's theory of diffusion of innovations, was elaborated and validated to specifically evaluate this phenomenon. Preliminary results show that the translation of training into practice might be suboptimal. The validated questionnaire can now be used to fully understand the set of factors influencing this new practice.

Cross-cultural adaptation, reliability, internal consistency and validation of the Spinal Function Sort (SFS) for French- and German-speaking patients with back complaints.

Introduction Functional subjective evaluation through questionnaire is fundamental, but not often realized in patients with back complaints, lacking validated tools. The Spinal Function Sort (SFS) was only validated in English. We aimed to translate, adapt and validate the French (SFS-F) and German (SFS-G) versions of the SFS. Methods Three hundred and forty-four patients, experiencing various back complaints, were recruited in a French (n = 87) and a German-speaking (n = 257) center. Construct validity was estimated via correlations with SF-36 physical and mental scales, pain intensity and hospital anxiety and depression scales (HADS). Scale homogeneities were assessed by Cronbach's α. Test-retest reliability was assessed on 65 additional patients using intraclass correlation (IC). Results For the French and German translations, respectively, α were 0.98 and 0.98; IC 0.98 (95% CI: [0.97; 1.00]) and 0.94 (0.90; 0.98). Correlations with physical functioning were 0.63 (0.48; 0.74) and 0.67 (0.59; 0.73); with physical summary 0.60 (0.44; 0.72) and 0.52 (0.43; 0.61); with pain -0.33 (-0.51; -0.13) and -0.51 (-0.60; -0.42); with mental health -0.08 (-0.29; 0.14) and 0.25 (0.13; 0.36); with mental summary 0.01 (-0.21; 0.23) and 0.28 (0.16; 0.39); with depression -0.26 (-0.45; -0.05) and -0.42 (-0.52; -0.32); with anxiety -0.17 (-0.37; -0.04) and -0.45 (-0.54; -0.35). Conclusions Reliability was excellent for both languages. Convergent validity was good with SF-36 physical scales, moderate with VAS pain. Divergent validity was low with SF-36 mental scales in both translated versions and with HADS for the SFS-F (moderate in SFS-G). Both versions seem to be valid and reliable for evaluating perceived functional capacity in patients with back complaints.

Validation of real-time methylation-specific PCR to determine O6-methylguanine-DNA methyltransferase gene promoter methylation in glioma.

Epigenetic silencing of the DNA repair protein O(6)-methylguanine-DNA methyltransferase (MGMT) by promoter methylation predicts successful alkylating agent therapy, such as with temozolomide, in glioblastoma patients. Stratified therapy assignment of patients in prospective clinical trials according to tumor MGMT status requires a standardized diagnostic test, suitable for high-throughput analysis of small amounts of formalin-fixed, paraffin-embedded tumor tissue. A direct, real-time methylation-specific PCR (MSP) assay was developed to determine methylation status of the MGMT gene promoter. Assay specificity was obtained by selective amplification of methylated DNA sequences of sodium bisulfite-modified DNA. The copy number of the methylated MGMT promoter, normalized to the beta-actin gene, provides a quantitative test result. We analyzed 134 clinical glioma samples, comparing the new test with the previously validated nested gel-based MSP assay, which yields a binary readout. A cut-off value for the MGMT methylation status was suggested by fitting a bimodal normal mixture model to the real-time results, supporting the hypothesis that there are two distinct populations within the test samples. Comparison of the tests showed high concordance of the results (82/91 [90%]; Cohen's kappa = 0.80; 95% confidence interval, 0.82-0.95). The direct, real-time MSP assay was highly reproducible (Pearson correlation 0.996) and showed valid test results for 93% (125/134) of samples compared with 75% (94/125) for the nested, gel-based MSP assay. This high-throughput test provides an important pharmacogenomic tool for individualized management of alkylating agent chemotherapy.