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[Experimental and clinical determination of renal size after intravenous and intra-arterial administration of ionic and non-ionic contrast media].

A quantitative comparison of changes in renal size after the intravenous injection of contrast media showed no difference between ionic and non-ionic contrast medium. Contrast media of similar osmolarity (Metrizamide and Ioxaglate with 470 and 490 mosn/kg H2O) did show significant variations. Evidently properties of the contrast medium, other that their osmotic behaviour, are of significance (chemical structure, viscosity, iodine content, specific toxicity, and the effect on the permeability of biological membranes, calcium-binding activity, effect on coagulation). Changes in renal size could be reproduced by repeating the contrast injection at 24 to 48 hours.

Animals↗

Construction and screening of a genomic library specific for mouse chromosome 16.

We have established a protocol for producing libraries of specific mouse chromosomes. The mouse DNA-containing clones from a genomic library of a hamster-mouse somatic cell hybrid containing only one mouse chromosome are identified by screening with radiolabeled mouse repetitive sequences after specifically blocking hamster repetitive sequences; 95% of the mouse DNA-containing clones are identified. We have applied this protocol in producing a library of mouse chromosome 16, consisting of 14,200 clones or two "chromosome equivalents." Each clone occupies an individual well in a microtiter tray, allowing the entire library to be repeatedly and reproducibly plated and analyzed by hybridization. Further, we have established a protocol for making cDNA probes specifically depleted of highly repetitive sequences for probing libraries of genomic clones. By screening the chromosome 16 library with cDNA probes from mouse liver and brain, we demonstrate the feasibility of identifying expressed sequences and characterizing their patterns of expression. Such chromosome-specific libraries can facilitate the isolation of defined genetic loci as well as form the basis for the production of integrated transcriptional, genetic, and physical maps of entire chromosomes.

Animals↗

Determination of digestible energy values and fermentabilities of dietary fibre supplements: a European interlaboratory study in vivo.

The performance of methods to determine energy conversion factors for dietary fibre (DF) supplements and fermentability (D) values of their non-starch polysaccharides (NSP) was investigated. Heats of combustion, digestible energy (DE) and D values were determined on five DF supplements in five European laboratories on five separate occasions. In each instance the DF supplements were fed to juvenile male Wistar rats at two doses, 50 and 100 g/kg basal diet, for 3 weeks with food and faeces collected in the 3rd week. Among-laboratory variations in heats of combustion (delta Hc) were < 2%. DE values (kJ/g dry weight) at the upper and lower doses respectively were: 10.4 and 9.9 for a high-methoxyl apple pectin, 9.5 and 9.4 for a sugar-beet DF supplement, 12.2 and 12.7 for soyabean DF supplement, 3.8 and 4.0 for maize bran, and 0.3 and 0.3 for Solka-floc cellulose. Variations among laboratories, among occasions and among animals were < 1, < 2 and < 2.5 kJ/g respectively. The among-occasion: among-laboratory variance ratio for DE was 0.5, suggesting the method performed equally well in all laboratories. There was no evidence of learning of fatigue or fatigue in the performance of the method. D values were also independent of dose and at the high and lower doses were: pectin 0.92 and 0.95, sugar-beet NSP 0.68 and 0.68, soyabean NSP 0.86 and 0.88, maize bran 0.17 and 0.18, cellulose 0.07 and 0.06. Among-laboratory variance tended to increase with decreasing fermentability and ranged from 0.03 to 0.18. The DE and D data were not significantly different from a previously proposed relationship DE = 0.7 x delta Hc x D, where delta Hc is the heat of combustion of the supplement. We conclude that while the among-laboratory variation in the D of difficult-to-ferment NSP is too large for the reliable prediction of energy value the method for the direction determination of DE is both reproducible and repeatable, that DE is independent of dosage of DF supplement up to 100 g/kg diet, and that it is safe to discriminate between energy values with a precision of 3 kJ/g. The conversion of both DE and D to net metabolizable energy for the purpose of food labelling, tables and databases is described.

Animals↗

Persistence of cellular and humoral response to synthetic peptides from defined Plasmodium falciparum antigens.

The cellular and humoral immune responses to synthetic peptides reproducing the repeat sequences of two major vaccine candidates (circumsporozoite protein and Pfl55/RESA) were investigated in two groups of African subjects according to the length of their stay outside endemic areas. The relation between the lymphoproliferative response and the antibody levels to these antigens was studied. The results confirm the existence of T-cell epitopes within the repeat sequences of the CS protein and the Pfl55/RESA capable of inducing lymphocyte proliferation. Cellular response to all studied peptides was more frequently observed in individuals living in France for less than one year than in individuals living in France for a longer time. T-cell proliferation in the presence of the tetrapeptide and of the octapeptide from the C-terminus repeat of Pfl55/RESA was related, with an immunodominance of the tetrapeptide over the octapeptide. Cellular responses to the CS protein repeat and to the 11-amino-acid peptide from the Pfl55/RESA N-terminus were the longest lasting after termination of exposure. In a given individual, cellular and humoral responses were not related for any peptide studied.

Adolescent↗

A method to estimate the uncertainty of measurements in a conglomerate of instruments/laboratories.

A comparison of the performance, at two concentrations, of the measurement procedures for 23 common components is presented. For each component the within-, pure between- and total variations have been calculated and an imprecision profile indicated. Thus a pragmatic, readily and generally applicable method is advised to describe the performance of laboratories under repeatability and reproducibility conditions. The calculated total variation is compared with the results estimated from the biological variation of the properties. In most cases the within-laboratory variation is better than that given by the reference database, whereas in about half of the procedures the total variation exceeds that of the reference database. The outcome illustrates the transferability problems that multicentre studies and the diagnosis of patients face by measuring samples using a conglomerate of instruments/laboratories that are not aligned. The calculation procedures are detailed in an appendix and the calculation and presentation of the results are made with a custom-made worksheet program.

Clinical Laboratory Techniques↗

Gas chromatographic determination of organochlorine pesticides in cow milk.

A gas chromatographic method of use in determining the following organochlorine pesticides in cow milk has been adapted and validated: alpha-hexachlorocyclohexane (alpha-HCH), beta-HCH, lindane, delta-HCH, hexachlorobenzene, aldrin, heptachlor, heptachlor epoxide, mirex, 2,4-dichlorodiphenyltrichloroethane (2,4-DDT), 4,4-DDT, 2,4-dichlorodiphenyldichloroethane (2,4-DDD), 4,4-DDD, 2,4-dichlorodiphenylethane (2,4-DDE) and 4,4-DDE. The method studied consists of three steps: the extraction of fat by centrifugation, mixing with anhydrous sodium sulfate, and transfer to a glass column and elution with hexane; purification of the extract via concentrated sulfuric acid treatment; and determination of the organochlorine pesticides by gas chromatography equipped with an electron capture detector. The analytical parameters determined to estimate the quality of the method yielded the following results: correlation coefficients (r) of the calibration curves were greater than 0.996, the variation coefficients of the response factors were less than 4.90%--with the exception of aldrin (5.53%) and lindane (5.75%)--the relative standard deviation of the slope (Sm rel) being < or =1.52%. Zero was included in the confidence limits of the independent term range. The method repeatability and reproducibility values were < or =4.73% and < or =5.79%, respectively. The recovery values of the pesticides analyzed were in the range of 81.92% (alpha-HCH) to 105.49% (4,4-DDT), except in the case of aldrin and heptachlor epoxide, which yielded recovery values of under 40%. The quantification limits of the method were < or =1 ng/g, with the exception of mirex (5 ng/g).

Animals↗

Measurement of the response time of an electrosensitive protective device in the process of its certification.

In the process of testing an electrosensitive protective system, determining its response time is of crucial importance. A unique double penetration method of measuring electrosensitive protective device (ESPD) response time has been worked out in the Central Institute for Labour Protection. In the first step, low speed penetration enables the detection zone border to be localised. In the second step of measurement, the probe is injected at a high speed and response time is measured. Three different ways have been taken for validation of the method: theoretical analysis, calibration of the stand, and taking a series of measurements. The double penetration method enables ESPD response time measurement results to be obtained, the accuracy, repeatability, and reproducibility of which is satisfactory enough to be assessed objectively.

Accidents, Occupational↗

Development, validation and characterization of an analytical method for the quantification of hydrolysable urinary metabolites and plasma protein adducts of 2,4- and 2,6-toluene diisocyanate, 1,5-naphthalene diisocyanate and 4,4'-methylenediphenyl diisocyanate.

Occupational exposure to diisocyanates within the plastic industry causes irritation and disorders in the airway. The aim of this study was to develop, validate and characterize a method for the determination of 2,4-toluenediamine (2,4-TDA), 2,6-toluenediamine (2,6-TDA), 1,5-diaminonaphthalene (1,5-NDA) and 4,4'-methylenedianiline (4,4'-MDA) in hydrolysed urine and plasma, and to study the correlation between the plasma and urinary levels of these potential biomarkers of 2,4-toluene diisocyanate (2,4-TDI), 2,6-toluene diisocyanate (2,6-TDI), 1,5-naphthalene diisocyanate (1,5-NDI) and 4,4'-methylenediphenyl diisocyanate (4,4'-MDI), respectively. Samples were hydrolysed with 0.3 M NaOH at 100 degrees C for 24 h. The diamines were extracted, derivatized with pentafluoropropionic acid anhydride, and quantified by selected ion monitoring on gas chromatography-mass spectrometry. The repeatability and reproducibility of the method were 7-18% and 7-19%, respectively. Dialysis experiments showed that the metabolites of 2,4-TDI, 2,6-TDI, 1,5-NDI and 4,4'-MDI in plasma were exclusively protein adducts. No free diamines were found in urine, indicating that all diisocyanate-related metabolites were in a conjugated form. For each diisocyanate-related biomarker, there were strongly significant correlations (p<0.001) between individual levels of metabolites in plasma and urine, with Spearman's rank correlation coefficient (rs) values of 0.74-0.90. The methods presented here will be valuable for the development of biological monitoring methods for diisocyanates.

Biomarkers↗

Test-retest intra-rater reliability of grip force in patients with stroke.

OBJECTIVE: Coefficients of repeatability and reproducibility can be guides in differentiating between real changes and measurement error. The aim was to evaluate test-retest intra-rater reliability of a clinical procedure measuring grip force with Grippit in stroke patients, to assess relationship between grip force of the hands and between sustained and peak grip force. PATIENTS AND METHODS: Eighteen patients were tested using the Grippit at two occasions one hour apart. Each occasion comprised three consecutive trials per hand. RESULTS: The paretic hand needs to score a 50 N change within and between occasions to exceed the measurement error in 95% of the observations, irrespective of calculation method. Expressed by CV(within) the measurement error was 10%. There was no learning or fatigue effect during measuring. There was a wide variation between subjects but the mean ratio between sides was 0.66. The mean ratio between sustained and peak grip force was 0.80-0.84. CONCLUSION: The measurement errors were acceptable and the instrument can be recommended for the use in stroke patients at a department of rehabilitation medicine.

Female↗

Equivalence in test assay method comparisons for the repeated-measure, matched-pair design in medical device studies: statistical considerations.

In medical device clinical studies, including therapeutic device and in vitro test assay method studies, the investigator is frequently interested in demonstrating the equivalence of clinical response, generally in continuous measurements, between a standard assay method and a new assay method over various occasions or times. The new assay method may be less invasive or more convenient or cheaper to use than the standard assay method. In this paper, several statistical approaches are discussed, including various repeated-measure regression models, the simultaneous 95% confidence interval for paired mean differences derived from Hotelling's multivariate T2 analysis for repeated-measure, paired data, repeatability and reproducibility studies, and concordance correlation coefficient.

Confidence Intervals↗

Investigation into the thermal distribution of microwave helical antennas designed for the treatment of Barrett's oesophagus.

A set of helical microwave antennas was designed to investigate their potential use in thermal therapy of Barrett's oesophagus. The antennas had a diameter of up to 3.3 mm and various lengths between 20 and 37 mm; these were designed to operate at 915 MHz. Sets of polytetrafluoroethylene (PTFE) formers were constructed to improve the repeatability and reproducibility of the helix manufacture. Small diameter copper wire was wound over the formers and connected to the coaxial cable at the inner and outer conductor junctions. The power deposition profiles of the antennas were measured in a muscle-equivalent phantom using an infrared camera. The effects of antenna length and coil spacing were characterized. It was observed that uniform temperature profiles along the antenna length were achieved with a length of wire of 99 mm +/- 2 mm. The effective heating length (length of the antenna that exhibits > 50% of the maximum temperature rise) was comparable to the antenna length. The radial penetration depth of 50% of the antenna surface temperature for the optimum 20 mm antenna was 2.5 mm from the antenna outer surface.

Barrett Esophagus↗

A simple and rapid method for determination of G gamma:A gamma globin chain synthetic ratio.

A simple and rapid method for the determination of the G gamma- to A gamma-globin chain relative synthetic ratios is described. This technic uses 35S-methionine as the labeled amino acid and introduces a simple procedure for Hb-F purification. The chain separation is based on slab gel isoelectric focusing, and the gel after visualization of protein bands without staining is cut and solubilized in periodic acid, and the radioactivity is counted. This method can be applied in blood samples with low Hb-F levels, and as many as 20 samples can be analyzed at one time. The method was found to yield the expected synthetic ratio, and its reproducibility and repeatability were found to be high.

Fetal Blood↗

Differences in the recovery of excitability in sensory and motor axons of human median nerve.

Following conduction of an action potential there is a stereotyped sequence of changes in excitability as axons are initially refractory, then superexcitable and finally subexcitable. These activity-dependent oscillations in excitability subside over 100 ms and together constitute the recovery cycle. The present study was undertaken first to document the recovery cycle of sensory and motor axons of different threshold and, secondly, to compare the changes in sensory axons with those in motor axons. A computerized threshold-tracking system was used to measure recovery cycles in six healthy subjects; stimuli were applied to the median nerve at the wrist. Changes in the threshold required to produce an antidromic compound sensory action potential (CSAP) and an orthodromic compound muscle action potential (CMAP) of fixed amplitude (30%, 50% and 70% of maximal) were recorded following a single supramaximal conditioning stimulus. Normalized recovery cycles were identical for axons of different threshold, whether sensory or motor, and were reproducible on repeat testing. However, there were significant differences between the changes in sensory and motor axons, with greater supernormality and greater late subnormality in motor axons. The greater changes in motor axons could not be explained by differences in the strength-duration properties of sensory and motor axons. There are biophysical differences in the properties of sensory and motor axons and these differences may underlie the differential susceptibility of sensory and motor axons in peripheral nerve disorders.

Adult↗

Spatial representation of odours in the antennal lobe of the moth Spodoptera littoralis (Lepidoptera: Noctuidae).

Glomeruli within the antennal lobe (AL) of moths are convergence sites for a large number of olfactory receptor neurons (ORNs). The ORNs target single glomeruli. In the male-specific cluster of glomeruli, the macroglomerular complex (MGC), the input is chemotypic in that each glomerulus of the MGC receives information about a specific component of the conspecific female sex pheromone. Little is known about how neurons that detect other odorants arborize in and amongst glomeruli. The present study focuses on how sex pheromones and biologically relevant semiochemicals are represented in the ALs of both sexes of the moth Spodoptera littoralis. To assess this, we optically measured odour-evoked changes of calcium concentration in the ALs. Foci of calcium increase corresponded in size and shape with anatomical glomeruli. More than one glomerulus was normally activated by a specific non-pheromonal odorant and the same glomerulus was activated by several odorants. All odorants and pheromone components tested evoked unique patterns of glomerular activity that were highly reproducible at repeated stimulations within an individual. Odour-evoked patterns were similar between individuals for a given odorant, implicating a spatial olfactory code. In addition, we demonstrated that activity patterns evoked by host-plant related volatiles are similar between males and females.

Animals↗

Determination of mangiferin, jateorrhizine, palmatine, berberine, cinnamic acid, and cinnamaldehyde in the traditional Chinese medicinal preparation Zi-Shen pill by high-performance liquid chromatography.

High-performance liquid chromatography is employed to determine the contents of six marker components such as mangiferin, jateorrhizine, palmatine, berberine, cinnamic acid, and cinnamaldehyde in the traditional Chinese medicinal preparation Zi-Shen pill. The separation is performed on a C(18) column by stepwise gradient elution with water (0.2%, v/v, triethylamine adjusted to pH 4 with phosphoric acid)-methanol-acetonitrile (0.01 min, 98:0:2; 20 min, 80:5:15; 30 min, 65:13:22; and 55 min, 65:13:22) as the mobile phase at a flow rate of 0.9 mL/min, with UV detection at 280 nm. Six regression equations show good linear relationships between the peak area of each marker and concentration. The recoveries of the markers listed are 95.5%, 98.3%, 96.8%, 99.5%, 101.7%, and 102.1%, respectively. The repeatability and reproducibility (relative standard deviation) of the method are less than 2.5% and 3.3%, respectively.

Acrolein↗

Determination of safflor yellow A, puerarin, ferulic acid, ginsenoside Rg1, and Rb1 in the Traditional Chinese Medicinal preparation Naodesheng injection by high-performance liquid chromatography.

High-performance liquid chromatography is employed to determine the contents of five mark components, safflor yellow A, puerarin, ferulic acid, ginsenoside Rg1, and Rb1, in the Traditional Chinese Medicinal preparation Naodesheng injection. The separation is performed on a C18 column by stepwise gradient elution with water (0.1%, v/v, phosphoric acid)-acetonitrile (0 min, 86:14; 48 min, 75:25; and 68 min, 50:50) as the mobile phase at a flow rate of 1.0 mL/min, with UV detection at 203 nm. Five regression equations show a good linear relationship between the peak area of each marker and concentration. The recoveries of the markers listed are 99.6%, 100.2%, 99.7%, 100.0%, and 99.7%, respectively. The repeatability and reproducibility (relative standard deviation) of the method are less than 1.4% and 1.8%, respectively.

Chalcone↗

Effects of adenosine on atrial refractoriness and arrhythmias.

OBJECTIVE: Transient atrial fibrillation is sometimes observed following adenosine administration and adenosine is known to shorten atrial action potential duration and refractory period. This study was designed to characterise the dose-response relationship of adenosine on these variables relative to arrhythmia induction with single atrial premature stimuli. The effects of adenosine during sustained atrial flutter were also determined. METHODS: Intravenous bolus doses of adenosine were given to pentobarbitone anaesthetised dogs following cervical vagotomy and autonomic blockade with atropine and nadolol. Monophasic action potential catheter recordings were obtained from the right atrium and a programmable stimulator was used for pacing. RESULTS: Placebo had no effect on monophasic action potential duration (MAPD) or atrial effective refractory period (ERP) and no arrhythmias were observed. Adenosine (0.1-1.0 mg.kg-1) produced dose related decreases in MAPD and ERP and transient atrial fibrillation (5-122 s) was repeatedly and reproducibly induced in 12 dogs. In six dogs, intravenous dipyridamole (0.25 mg.kg-1) enhanced the effects of adenosine on MAPD and ERP and increased the incidence of atrial fibrillation. In another six dogs, 8-sulphophenyltheophylline (5.0 mg.kg-1, intravenously) markedly blunted the effects of adenosine, and atrial fibrillation could no longer be induced by premature stimuli. In a separate series of experiments the effects of adenosine were evaluated in seven dogs in which sustained atrial flutter could reproducibly be induced by rapid atrial pacing. Administration of placebo never caused termination of the arrhythmia, whereas intravenous boluses of adenosine (0.1-1.0 mg.kg-1) decreased and produced variation in atrial flutter cycle length, and then terminated the arrhythmia in all cases. These effects of adenosine were also enhanced by dipyridamole and antagonised by 8-sulphophenyltheophylline. CONCLUSIONS: Adenosine produces a receptor mediated shortening of monophasic action potential duration and refractoriness which increases vulnerability to transient atrial arrhythmias. During sustained atrial flutter, these effects may also contribute to destabilisation and termination of the arrhythmia.

Action Potentials↗

Muscarinic agonist-induced actions on potassium and calcium channels in atrial myocytes: differential desensitization.

Desensitization to the effects of acetylcholine (ACh) or carbachol (CCh) on ACh-regulated potassium current (IK(ACh)) and the voltage-dependent L-type, calcium current (ICa(L)) was studied in single guinea-pig atrial cells under voltage clamp at room temperature (22-24 degrees C). At a holding potential of -40 mV, CCh (100 microM) activated IK(ACh) repeatedly and reproducibly when applied for 15-s periods with 60-s intervals between applications. Prolonged (10 min) exposure to CCh caused a time-dependent decline of IK(ACh) and a marked reduction (desensitization) of the response to subsequent application of CCh. In the absence of guanosine triphosphate (GTP) in the pipette, only partial resensitization occurred within 20 min after CCh washout. The desensitization to CCh could also be obtained when adenosine (20 microM) was used to activate this current. Therefore, this desensitization must be heterologous. In the presence of isoproterenol (ISO, 3 microM), ICa(L) increased. Acetylcholine continued to inhibit this current at a time when its activation of IK(ACh) had become desensitized. Muscarinic inhibition of ICa(L) eventually desensitized but only after IK(ACh) desensitization had occurred. Because of its heterologous nature, as well as of the ability of ACh to desensitize in the presence of intrapipette dextran, which is reported to inhibit beta-adrenergic receptor kinase (beta ARK), it seems unlikely that beta ARK-dependent phosphorylation of the muscarinic receptor (mAChR) accounted for desensitization. The differential desensitization time course is consistent with the hypothesis that different subunits of Gi, the inhibitory guanine nucleotide binding protein, not only transduce the agonist effects of ACh on IK(ACh) and ICa(L) but are also subjected to different inactivation/regulation processes.

Acetylcholine↗