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Distribution of human SNPs and its effect on high-throughput genotyping.

Utilizing the results of extensive single nucleotide polymorphism (SNP) studies in humans, stimulated by the International HapMap Project, we present evidence that SNPs are not randomly spaced across the genome, but are somewhat clustered. This observation has important consequences for assay design, since hidden variants in primer sites can affect the accuracy of data. Indeed, using data from the calibration exercises of the HapMap Project, we found instances in which primer site mutations caused allele dropout and other genotyping failures. Given the dynamic nature of SNP discovery, it was inevitable that SNPs would be identified in the primer sites of many assays used for HapMap genotyping. We found that assays with such primer site mutations were correlated with elevated rates of genotype failure and allele dropout. This suggests that taking nearby SNPs into account is important for optimal genotyping assay design.

Alleles↗

Screening for and validated quantification of amphetamines and of amphetamine- and piperazine-derived designer drugs in human blood plasma by gas chromatography/mass spectrometry.

The classical stimulants amphetamine, methamphetamine, ethylamphetamine and the amphetamine-derived designer drugs MDA, MDMA ('ecstasy'), MDEA, BDB and MBDB have been widely abused for a relatively long time. In recent years, a number of newer designer drugs have entered the illicit drug market. 4-Methylthioamphetamine (MTA), p-methoxyamphetamine (PMA) and p-methoxymethamphetamine (PMMA) are also derived from amphetamine. Other designer drugs are derived from piperazine, such as benzylpiperazine (BZP), methylenedioxybenzylpiperazine (MDBP), trifluoromethylphenylpiperazine (TFMPP), m-chlorophenylpiperazine (mCPP) and p-methoxyphenylpiperazine (MeOPP). A number of severe or even fatal intoxications involving these newer substances, especially PMA, have been reported. This paper describes a method for screening for and simultaneous quantification of the above-mentioned compounds and the metabolites p-hydroxyamphetamine and p-hydroxymethamphetamine (pholedrine) in human blood plasma. The analytes were analyzed by gas chromatography/mass spectrometry in the selected-ion monitoring mode after mixed-mode solid-phase extraction (HCX) and derivatization with heptafluorobutyric anhydride. The method was fully validated according to international guidelines. It was linear from 5 to 1000 micro g l(-1) for all analytes. Data for accuracy and precision were within required limits with the exception of those for MDBP. The limit of quantification was 5 micro g l(-1) for all analytes. The applicability of the assay was proven by analysis of authentic plasma samples and of a certified reference sample. This procedure should also be suitable for confirmation of immunoassay results positive for amphetamines and/or designer drugs of the ecstasy type.

Amphetamines↗

Screening for and validated quantification of phenethylamine-type designer drugs and mescaline in human blood plasma by gas chromatography/mass spectrometry.

In recent years, several newer designer drugs of the so-called 2C series such as 2C-D, 2C-E, 2C-P, 2C-B, 2C-I, 2C-T-2, and 2C-T-7 have entered the illicit drug market as recreational drugs. Some fatal intoxications involving 2C-T-7 have been reported. Only scarce data have been published about analyses of these substances in human blood and/or plasma. This paper describes a method for screening and simultaneous quantification of the above-mentioned compounds and their analog mescaline in human blood plasma. The analytes were analyzed by gas chromatography/mass spectrometry in the selected-ion monitoring mode, after mixed-mode solid-phase extraction (HCX) and derivatization with heptafluorobutyric anhydride. The method was fully validated according to international guidelines. Validation data for 2C-T-2 and 2C-T-7 were unacceptable. For all other analytes, the method was linear from 5 to 500 microg/L and the data for accuracy (bias) and precision (coefficient of variation) were within the acceptance limits of +/-15% and <15%, respectively (within +/-20% and <20% near the limit of quantification of 5 microg/L).

Designer Drugs↗

Discrepant karyotypes after second- and third-trimester combined placentacentesis/amniocentesis.

Cytogenetic data are presented from a total of 1306 consecutive pregnancies with successful diagnosis obtained from both chorionic villi after short-time culture (CVS-SC) and amniotic fluid cell cultures (AC); samples had been taken simultaneously at combined placentacentesis (placental biopsy) and amniocentesis during the second (92.8 per cent) and third (7.2 per cent) trimesters. Concordant results were obtained in 1218 pregnancies with a normal karyotype and in 62 pregnancies with an aberrant fetal karyotype. Discrepant, i.e. false-positive and false-negative, results were found in 26 cases (2 per cent). From these data the accuracy of CVS-SC, defined as the proportion of all correct diagnoses, is calculated to be 98 per cent. Three non-mosaic and 14 mosaic false-positive results obtained after CVS-SC could not be confirmed by AC. Related to 1235 true normal fetal karyotypes, the specificity of CVS-SC, i.e. the proportion of normal karyotypes correctly diagnosed, amounts to 98.6 per cent. In nine pregnancies, an aberrant fetal karyotype detected after AC was missed by CVS-SC. The sensitivity of CVS-SC, i.e. the proportion of abnormal fetuses correctly diagnosed (62 out of 71), amounts to 87.3 per cent in our study group.

Amniocentesis↗

An examination of the efficiency of some quality assurance methods commonly employed in clinical trials.

The cost and efficiency of training clinical centre staff and of duplicate data entry in clinical trials is reviewed. Training is an essential component of quality assurance programmes and it is usually carried out at regular intervals in long-term clinical trials. Initial training of staff and regular retraining is important to assure standardization and it can lead to reduced trial costs. Interim training for new staff and for remedial purposes is less efficient than regularly scheduled training sessions. Regional centres for training and the use of computer aided instruction are two ways such interim training can be made more efficient and standardized. Duplicate data entry or verification can result in a substantial reduction in data entry errors depending on the nature of the data being keyed. Selective verification should especially be considered for important fields for which consistency checks cannot be performed, that are alphabetic or that are several characters in length. Quality assurance procedures should be implemented to monitor data entry accuracy in clinical trials.

Documentation↗

A data-analytic approach for estimating lead time and screening benefit based on survival curves in randomized cancer screening trials.

Screening tests are used frequently for control of diseases such as cancer. The increased survival time of screen-detected cases over those that are detected clinically may be due in part to 'lead time', or the length of time by which the disease is diagnosed earlier by screening in the presence or absence of any real extension in survival time. A realistic evaluation of screening needs to assess the true benefit of screening; that is, the length of time by which survival has been extended, beyond merely the time of the advanced diagnosis. The comparison of survival measured from time of entry between cases in a screening arm and in a control arm in randomized studies avoids the lead time bias. If the effects of average lead time and average benefit on survival are additive, these effects can be estimated by recognizing that (a) the difference in survival curves since time of diagnosis confounds benefit and lead time, but (b) the difference in survival curves since time of start of study involves benefit only. The method is evaluated on simulated data for its accuracy and may be used on data from randomized studies.

Bias↗

A microtitre plate method for measuring biopterin with cryopreserved Crithidia fasciculata.

The assay of biopterin derivatives in dried blood spots is used by us in initial screening for inherited defects in tetrahydrobiopterin synthesis. The previously described method (1) required aseptic technique and microbiological facilities. The modification detailed here has the advantages of antibiotic cover, which overcomes these needs and microtitre plate technology allowing the incubation time to be halved with precision and accuracy retained. Data reduction facilities may be applied.

Adult↗

Practical experience and issues in designing and performing population pharmacokinetic/pharmacodynamic studies.

An expert meeting to discuss issues relating to the design of population pharmacokinetic/pharmacodynamic (PK/PD) studies was held in Brussels in March 1995, under the auspices of the European Co-operation in Science and Technology (COST), Medicine (B1) programme. The purpose of the meeting was to discuss the experts' experience in designing and performing population PK/PD studies. The topics discussed were current practice, logistical issues, ensuring the accuracy of data, covariate assessment, communication, and protocol design.

Belgium↗

Facts on optic flow.

We employ an optimal solution to both the "shape from motion problem" and the related problem of the estimation of self-movement on a purely optical basis to deduce practical rules of thumb for the limits of the optic flow information content in the presence of perturbation of the motion parallax field. The results are illustrated and verified by means of a computer simulation. The results allow estimates of the accuracy of depth and egomotion estimates as a function of the accuracy of data sampling and the width of field of view, as well as estimates of the interaction between rotational and translational components of the movement.

Humans↗

A three-dimensional photographic method for documentation and measurement of dental conditions.

A 3-D photographic technique was developed for the registration of bite conditions. This technique may replace plaster models for archiving purposes. To achieve this, photographic equipment and computer programs should be user friendly, not too expensive, and allow measurements that satisfy scientific demands. This purpose was fulfilled by combining the following elements: a photographic unit with standardized conditions for 3-D photography and a reference object with known coordinates where bite impressions or models are placed during photography. To reconstruct 3-D coordinates from photographs these are placed on a digitizer connected to a computer programmed for reconstructing 3-D coordinates from the digitized 2-D data. The accuracy of the 3-D reconstruction of coordinates, x, y, z, was tested by means of 3-D photographs of a test object with reference points having a precision of +/- 0.5 micron. The mean error for distances of 5 mm and 15 mm varied between -0.17 mm and +0.15 mm. Measurements of 45 degrees and 90 degrees angles had an accuracy that varied from -0.5 degree to +0.9 degree and from -1.2 degrees to +1.5 degrees respectively. An angle between lines and a plane of 54.7 degrees displayed a mean error of +/- 1.6 degrees. The test of the accuracy of the 3-D photographic technique for calculating distances and angles demonstrated that it fulfills demands for scientific applications on clinical material.

Calibration↗

Evaluation of the Cobas Bact automated system for susceptibility testing of Enterobacteriaceae, Pseudomonas aeruginosa, and Enterococcus faecalis to azlocillin, mezlocillin, and ciprofloxacin compared to NCCLS and DIN standards.

The aim of automated susceptibility testing systems like the Cobas Bact is to provide the clinicians with rapid and reliable results for the care of patients and to decrease the work load in microbiological laboratories. Because data about accuracy on mezlocillin, azlocillin and ciprofloxacin were lacking, we investigated 184 bacterial strains and compared the results of the Cobas Bact susceptibility testing to standardized agar dilution and agar diffusion methods. Essential correlations for all methods compared exceeded 90% for the three chemotherapeutics and all species investigated, with the exception of Pseudomonas aeruginosa. On an average only 1.5% very major errors were observed with the several species of Enterobacteriaceae, whereas P. aeruginosa and Enterococcus faecalis were characterized by the complete absence of very major errors when Cobas Bact was correlated to NCCLS agar diffusion.

Autoanalysis↗

Measurement of CO2 response with the breath-by-breath automatic acquisition of the breathing pattern and occlusion pressure.

OBJECTIVE: Our objective is to present a methodology for the automated acquisition and storage of BP and P0.1 during a CO2 rebreathing test. METHODS: The system consists of a microcomputer with additional circuits and an automatic electronically controlled valve to occlude the inspiratory airway. Data collection and data processing are separate programs. Airway pressure and flow are digitized at a 100-Hz rate, while PETCO2 is determined and P0.1 is measured on a breath-by-breath basis. Off-line processing calculates the BP variables, generates a correlation matrix (VE/PETCO2, TTOT/PETCO2, TI/PETCO2, TE/PETCO2, [VT/TI]/PETCO2, [TI/TTOT]/PETCO2, P0.1/PETCO2), and edits graphic data. The accuracy of the volume and pressure measurements was tested by comparing known volumes provided by a syringe (n = 100) and a series of pressures controlled by a water manometer (n = 41) on the one hand, with volumes and pressures measured by the device. The accuracy of the time intervals and P0.1 was assessed by comparing in 10 healthy subjects the values measured manually on a graphic recording with those provided by the device (n = 170). RESULTS: Volumes:Vmeasured = 0.99 x Vcontrolled, r = 0.99, p < 0.001. Pressures:Pmeasured = 0.97 x Pcontrolled + 0.09, r = 0.98, p < 0.001. Inspiratory time:TIautomatic = 0.91 x TIgraphic + 0.22, r = 0.93, p < 0.001. Expiratory time:TEautomatic = 0.93 x TEgraphic + 0.34, r = 0.95, p < 0.001. Occlusion pressure:P0.1automatic = 0.95 x P0.1graphic + 0.62, r = 0.94, p < 0.001. Reproducibility was assumed to be represented by the intraindividual coefficient of variation of the CO2 response. The comparison of an automatic breath-to-breath method with a graphic manual recording revealed significantly less variability with the former (VE/PETCO2: 15.2 +/- 4.5% vs 22.5 +/- 6.3%, p < 0.01; P0.1/PETCO2:8.3 +/- 4.3% vs 19.7 +/- 7.2%, p < 0.001; [VT/TI]/PETCO2:9.1 +/- 3.5% vs 14.5 +/- 5.3%, p < 0.05). CONCLUSION: Our automated acquisition and storage of waveforms and breath-by-breath determination of BP and P0.1 provide an easy and thorough analysis of the respiratory response to CO2 and decrease the variability of the results.

Anesthesia, Closed-Circuit↗

Determination of iron and iron-binding capacity in serum without blank sample.

A highly sensitive monoreagent for the determination of iron and total iron-binding capacity (TIBC) in serum is described in this paper. It contains the dye chromazurol B which gives with iron a high molar absorption complex (A630nm = 1.6 X 10(5) l/mol cm). The more complex problem in the study of the reagent was to make negligible the interferences of the substances in the sera in order to get the procedure without blank samples or deproteinization. Until now none of the reagents presents these peculiarities, that might render easier both the manual analysis and the application to the automatic apparatus. Furthermore, the stability of the reagent (2 years) and the reliability of the results, as detected by data on accuracy, precision (coefficient of variation = 1.9%), linearity (up to 110 mumol/l) and correlation with three different methods, indicate that this procedure is particularly suitable for routine clinical chemistry.

Benzoates↗

Concentration of LAS and boron in the Itter. Comparison of measured data with results obtained by simulation with the GREAT-ER software.

The GREAT-ER software was used to calculate linear alkylbenzene sulphonate [LAS] and boron concentrations in the Itter stream in North Rhine-Westfalia, Germany. The aim was to investigate the predictive strength of this newly developed tool and to compare its results to measured data. Substance-specific input data which were used in this scenario were partly generic (e.g. LAS and sodium perborate tetrahydrate consumption figures for Germany) and partly (site-)specific data (e.g. half-life time for LAS elimination). The comparison with the measured data reveals that the model predictions for LAS and boron are correct at least within a factor of two, only if generic German consumption data is applied. By using refined input data, the accuracy can be increased further.

Alkanesulfonic Acids↗

A stochastic control program to predict outcome and to support therapeutic decisions: a preliminary report.

BACKGROUND AND OBJECTIVES: Early noninvasive hemodynamic monitoring with an outcome predictor and a therapeutic decision support system may be useful to identify and correct hemodynamic deficiencies in emergency patients. The first aim was to apply a stochastic (probability) search and display model to predict outcome as early as possible. The second aim was to explore the usefulness of a therapeutic decision support system to evaluate the relative effectiveness of various therapies. METHODS: A stochastic control and display program based on noninvasive hemodynamic monitoring was applied in 100 consecutive critically ill patients admitted to the emergency department of an inner city public hospital. The program continuously displayed the noninvasive hemodynamic data and the patient's predicted survival probability (SP) that was based on the patient's diagnosis, covariates, and hemodynamic data. The accuracy of the SP at the initial resuscitation on admission to the emergency department (ED) was evaluated by the actual outcome at hospital discharge. The therapeutic decision support program evaluated the relative effectiveness of various therapies on based on their hemodynamic and SP responses and outcome of patients with similar clinical-hemodynamic states. RESULTS: The cardiac index, mean arterial pressure, arterial saturation, transcutaneous oxygen and carbon dioxide tensions were appreciably higher in survivors than in nonsurvivors in the initial resuscitation. Heart rate was higher in the nonsurvivors. The calculated Survival Probability (SP) of survivors averaged 81 +/- 1.4% in the first 24-hour observation period. It was 58 +/- 2.2% for nonsurvivors during this period. Misclassifications were 10/100 or 10%.

Adult↗

Merkel cell carcinoma: Is there a role for 2-deoxy-2-[f-18]fluoro-D-glucose-positron emission tomography/computed tomography?

PURPOSE: 2-Deoxy-2-[F-18]fluoro-D-glucose (FDG)-positron emission tomography (PET)/computed tomography (CT) is becoming widely available as a powerful imaging modality, combining the ability to detect active metabolic processes and their morphologic features in a single study. The role of FDG-PET/CT is proven in lymphoma, melanoma, colorectal carcinoma, and other cancers. However, there are rare malignancies such as Merkel cell carcinoma that can potentially be evaluated with PET/CT. We were therefore prompted to review our experience with FDG-PET/CT in the management of patients with Merkel cell carcinoma. PROCEDURES: This is a retrospective case series of six patients with Merkel cell carcinoma, 58-81 years old (average 69 +/- 8.3), who had whole-body PET/CT at our institution from January 1st, 2003 to August 31st, 2005. Two patients were women and four were men. Reinterpretation of the imaging studies for accuracy and data analysis from medical records were performed. RESULTS: Twelve examinations were acquired for the six patients (one patient had six PET/CT, one patient had two PET/CT, and four patients had one PET/CT). The injected FDG doses ranged 381.1-669.7 MBq (average 573.5 +/- 70.3). Four patients had the PET/CT as part of initial staging, and two patients had the exam for restaging (after surgery and XRT). A total of six Merkel lesions (pancreas, adrenal, lip, submandibular lymph nodes, cervical lymph nodes, and parapharyngeal soft tissue) were identified in three patients and confirmed on histopathological examination. The FDG uptake in these areas was intense, with maximum standardized uptake value (SUVmax) values of 5-14 (average 10.4 +/- 3.8). In one patient, the PET/CT scan identified abnormal focal distal sigmoid uptake that was biopsied and diagnosed as adenocarcinoma. Two patients had negative scans and had no clinical evidence of disease on follow-up office visits (up to one year after PET/CT). CONCLUSIONS: This case series suggests that FDG-PET/CT may have a promising role in the management of patients with Merkel cell carcinoma.

Aged↗

2-Deoxy-2-[F-18]fluoro-D-glucose positron emission tomography/computed tomography in the management of melanoma.

OBJECTIVES: 2-Deoxy-2-[F-18]fluoro-D-glucose (FDG)-positron emission tomography (PET)/computed tomography (CT) is widely available as a powerful imaging modality, combining the ability to detect active metabolic processes and their morphologic features in a single exam. The role of FDG-PET is proven in a variety of cancers, including melanoma, but the estimates of sensitivity and specificity are based in the majority of the published studies on dedicated PET, not PET/CT. Therefore, we were prompted to review our experience with FDG-PET/CT in the management of melanoma. METHODS: This is a retrospective study on 106 patients with melanoma (20-87 years old; average: 56.8 +/- 15.9), who had whole-body FDG-PET/CT at our institution from January 2003 to June 2005. Thirty-eight patients (35.9%) were women and 68 patients (64.1%) were men. Reinterpretation of the imaging studies for accuracy and data analysis from medical records were performed. RESULTS: All patients had the study for disease restaging. The primary tumor depth (Breslow's thickness) at initial diagnosis was available for 76 patients (71.7%) and ranged from 0.4 to 25 mm (average: 3.56 mm). The anatomic level of invasion in the skin (Clark's level) was determined for 70 patients (66%): 3, level II; 13, level III; 43, level IV; 11, level V. The administered dose of (18)F FDG ranged from 9.8 to 21.6 mCi (average: 15.4 +/- 1.8 mCi). FDG-PET/CT had a sensitivity of 89.3% [95% confidence interval (CI): 78.5-95] and a specificity of 88% (95% CI: 76.2-94.4) for melanoma detection. CONCLUSION: This study confirms the good results of FDG-PET/CT for residual/recurrent melanoma detection, as well as for distant metastases localization. PET/CT should be an integral part in evaluation of patients with high-risk melanoma, prior to selection of the most appropriate therapy.

Adult↗

On the time course and accuracy of spatial localization: basic data and a two-process model.

This article addresses the question how fast and accurate the location of a single stimulus can be perceived. In Experiment 1, we measured localization performance in a task which required subjects to perceive and report the location of a single target stimulus ('*' sign) presented in one square of an imaginary 25 x 19 grid. Two factors were varied: stimulus duration and stimulus eccentricity. Stimulus duration was manipulated by employing a backward masking stimulus. Ten intervals (stimulus onset asynchronies) separated target and masking stimulus: 25, 50, 75, 100, 125, 150, 200, 250, 300, and 350 ms. Stimulus eccentricity was manipulated by presenting the target stimulus at five different distances from the fixation point. The observer localized the target stimulus by moving the cursor from the middle of the grid (the initial fixation point) to the perceived target location by pressing the 'arrow' keys on the keyboard. Localization performance showed to be typically related to stimulus duration. That is, two components could be distinguished: The first component represented an initial steep rise in localization performance during the first 50 ms of stimulus duration; the second component represented a gradual rise in localization performance after 50 ms, reaching maximal performance at about 300 ms. We interpreted these two localization performance functions as reflecting the operation of two systems, namely the attentional system for the initial strong increase and the eye movement system for the subsequent gradual increase. In Experiment 2, we measured saccadic eye response latencies to clarify the role of eye movements in localization performance. It was found that in 98.4% of all trials saccades were executed, and, moreover, that saccadic eye response latency decreased with increasing stimulus duration. In Experiment 3, we compared localization performance in the absence and presence of eye movements and demonstrated that localization performance for stimulus durations up to 50 ms was independent of eye movements. Overall, the present findings were interpreted as evidence in support of a two-process model of localization performance in which a shift of attention is followed by a rapid eye movement toward the target location. In line with a continuous flow conception of visual information processing, our model assumes that location information takes time to develop in the visual system; hence, an observer's localization response may be based on qualitatively different processes operating on qualitatively different kinds of information. In case of short duration stimuli, information conveyed by transient cells is used by the attentional system to shift attention toward the target location; this results in course location information being available.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗