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Reproducibility and validity of dietary patterns assessed with a food-frequency questionnaire.

BACKGROUND: Recently, the analysis of dietary patterns has emerged as a possible approach to examining diet-disease relations. OBJECTIVE: We examined the reproducibility and validity of dietary patterns defined by factor analysis using dietary data collected with a food-frequency questionnaire (FFQ). DESIGN: We enrolled a subsample of men (n = 127) from the Health Professionals Follow-up Study in a diet-validation study in 1986. A 131-item FFQ was administered twice, 1 y apart, and two 1-wk diet records and blood samples were collected during this 1-y interval. RESULTS: Using factor analysis, we identified 2 major eating patterns, which were qualitatively similar across the 2 FFQs and the diet records. The first factor, the prudent dietary pattern, was characterized by a high intake of vegetables, fruit, legumes, whole grains, and fish and other seafood, whereas the second factor, the Western pattern, was characterized by a high intake of processed meat, red meat, butter, high-fat dairy products, eggs, and refined grains. The reliability correlations for the factor scores between the 2 FFQs were 0.70 for the prudent pattern and 0.67 for the Western pattern. The correlations (corrected for week-to-week variation in diet records) between the 2 FFQs and diet records ranged from 0.45 to 0.74 for the 2 patterns. In addition, the correlations between the factor scores and nutrient intakes and plasma concentrations of biomarkers were in the expected direction. CONCLUSIONS: These data indicate reasonable reproducibility and validity of the major dietary patterns defined by factor analysis with data from an FFQ.

Adult↗

[Morphometry of protuberant changes of the optic nerve head].

For objective evaluation of elevated changes in the optic nerve head, volumetric measurement of optic nerve heads in normal eyes, and in eyes with optic neuritis and papilledema was performed using a three-dimensional computerized digital image analyzer. To determine the reliability of the data, both intraphotographic and interphotographic error variance was studied. The reproducibility of the data was good, because the coefficient variation of elevated volume was small, 1.0 to 8.3% (mean; 4.5%) on intraphotographic study and 0.8 to 11.5% (mean; 6.5%) on interphotographic study in the optic neuritis and papilledema. The accuracy of the data was good enough for clinical use, presuming a change of less than 0.05 mm as measuring error.

Adult↗

Microbiology Galaxy Lab: The first community-driven gateway for reproducible and FAIR analysis of microbial data.

The explosion of microbial omics data has outpaced the ability of many researchers to analyze it, with complex tools and limited computational resources creating barriers to discovery. To address this gap, we present the Microbiology Galaxy Lab: a free, globally accessible, community-supported platform that combines state-of-the-art analytical power with user-friendly accessibility. Supported by the Galaxy and global microbiology communities, this platform integrates over 315 tool suites and 115 curated workflows, enabling comprehensive metabarcoding, (meta)genomic, (meta)transcriptomic, and (meta)proteomic data analysis within a FAIR-aligned environment. It also supports research in the health and infectious disease sectors, as well as in environmental microbiology. The platform's utility is exemplified through various use cases, including antimicrobial resistance tracking, biomarker prediction, microbiome classification, and functional annotation of key microbes. Built on reproducibility and community engagement, it supports creation, sharing, and updating of best-practice workflows. Over 35 tutorials and learning paths empower scientists, fostering an ecosystem that keeps resources at the forefront of microbial science. The Microbiology Galaxy Lab enables collective analysis, democratising research, thereby accelerating discovery across the global microbiology community (microbiology.usegalaxy.org, .eu, .org.au, .fr).

Journal Article↗

Processing and quality control of DNA array hybridization data.

MOTIVATION: The technology of hybridization to DNA arrays is used to obtain the expression levels of many different genes simultaneously. It enables searching for genes that are expressed specifically under certain conditions. However, the technology produces large amounts of data demanding computational methods for their analysis. It is necessary to find ways to compare data from different experiments and to consider the quality and reproducibility of the data. RESULTS: Data analyzed in this paper have been generated by hybridization of radioactively labeled targets to DNA arrays spotted on nylon membranes. We introduce methods to compare the intensity values of several hybridization experiments. This is essential to find differentially expressed genes or to do pattern analysis. We also discuss possibilities for quality control of the acquired data. AVAILABILITY: http://www.dkfz.de/tbi CONTACT: M.Vingron@dkfz-heidelberg.de

Animals↗

[Reproducibility of a self-administered questionnaire for dietary habits, smoking, and drinking].

Reproducibility of results from a self-administered questionnaire on dietary habits (the frequency of taking various foods and eating habits), smoking and drinking was examined to study the reliability of the questionnaire, stability of lifestyle, and the validity of the questionnaire in assessing lifestyle as a risk factor in chronic diseases. The study sample included 120 males and 173 females in a rural town in Kyoto Prefecture, who participated in a series of three health examinations in 1988, 1989, and 1992. A survey using the same questionnaire was performed before each health examination. Reproducibility was assessed by correlation (Spearman's rank correlation coefficient or kappa coefficient), exact agreement of category answered, and comparison of mean frequency. An attempt was made to separate reliability and stability from reproducibility using the data from the questionnaire obtained in the three years. Good reproducibility for one-year and four-year intervals was found for foods taken habitually or often (boiled rice, cooked rice gruel with tea, milk, coffee, fruits and bread), and habits (eating breakfast, some eating habits, smoking and drinking). The same was also found for cigarette or alcohol consumption among current consumers except that cigarette consumption over a four-year interval was more likely to change. The reliability of the questionnaire and stability of these items were considered satisfactory, suggesting that the questionnaire was a valid method of assessing long-term lifestyle. Poor reproducibility of results regarding consumption of green-yellow and other vegetables indicated both poor reliability and a change in consumption over four years, and suggested poor validity. However, disagreement in more than one category of vegetables was not large and misclassification in assessing them as risk factors and changes in frequency of consumption may not be large either. Although correlation coefficients for the items of dairy food and eating snacks were fair, disagreement in more than one category was relatively large which may have caused misclassification. The mean frequency of consumption of pickles and fishpaste, ham or sausage significantly decreased over the four year interval. This may reflect the influence of health education on reducing salt intake.

Adult↗

Cytotoxicity of precious and nonprecious alloys--experimental comparison of in vitro data from two laboratories.

The aim of this investigation was to evaluate and compare the reproducibility of cytotoxicity data generated in two different laboratories using the same testing protocols. A series of dental alloys that are widely used in both countries were chosen. These alloys (five precious, two nonprecious) were wet ground up to 1200 grit SiC, sterilized in 70% ethanol, and extracted in sterile culture medium for 7 days. Pure copper was used as a positive control and Teflonreg and media only were used as negative controls. Test and control samples were randomized and blinded to each laboratory. Cells, primary human gingival fibroblasts, and immortalized 3T3 fibroblasts, were exposed to the extracts for 24 h. Extract cytotoxicity was evaluated spectrophotometrically with the use of a mitochondrial enzyme activity assay. Data were collected from both laboratories, combined, and subjected to a mixed-model analysis of variance. No statistical difference was obtained for the immortalized 3T3 cells, except for two extracts in which differences between the two labs were significant but were still not cytotoxic. Furthermore, no statistical differences were found for the primary cells. These data strongly suggest that cytotoxicity tests performed in different laboratories with the use of the same test materials may lead to comparable results if sample preparation, cells, test procedures, and data analyses are carefully considered.

3T3 Cells↗

Some remarks on the calculation of the pore size distribution function of activated carbons.

Different authors investigated the effects of geometric and energetic heterogeneities on adsorption and on carbon characterization methods. In most theoretical studies carbon structure is modeled as parallel infinite graphite walls that form ideal slit-shaped pores of the fixed widths. In the literature there is the lack of systematic studies showing the influence of pore structural and Lennard-Jones (LJ) potential parameters on the pore-size distribution functions. Moreover, the parameters characterizing the properties of the adsorbed phase and the heterogeneity of the adsorbent surface should be taken into account. The Nguyen and Do method with proposed by us ASA algorithm, were utilized for the assessment of the porosity from the series of almost few thousands numerically generated local adsorption isotherms. The values of the mentioned-above parameters are varied over the wide range (ca. +/-20%) of the reference ones. Different types of the theoretical and experimental adsorption isotherms (nitrogen at 77 K) were taken into account as the global ones. They were related to the mechanism of the primary, secondary or mixed micropore filling. The variations in some above-mentioned parameters have significant effects only for PSDs (and for average pore widths) corresponding to the primary micropore filling mechanism. On the other hand, for the process of the secondary micropore filling, the influence of these parameters (without the BET coefficient for adsorption on a "flat" surface, c(s,B)) is rather insignificant. Nevertheless the differences between local and global adsorption isotherms (in the whole range of relative pressures) the absence of micropores having pore half width equal to ca. 1 nm on PSDs was observed for studied adsorbate-adsorbent systems with exceptions of the strictly microporous adsorbents and/or the low values of c(s,B). Comparison of the experimental data with the generated theoretical isosteric enthalpy of adsorption indicates that the phenomenal uptake observed from experiment can be explained in terms of the reasonable solid-fluid interaction parameters. Therefore, we varied the heterogeneity of the adsorbent surface via the strength and the range of the solid-fluid potential and the parameter c(s,B) in order to reproduce the experimental data of enthalpy of adsorption. Note that similar procedure was applied by Wang and Johnson to reproduce some hydrogen adsorption data measured for carbon nanofibres. The analysis of the obtained results shows that the selection of the values of the parameters of the intermolecular interactions and the quantities characterizing the properties of the adsorbed phase and the heterogeneity of the adsorbent walls for molecular simulations should be made with care and the influence of possible errors should be considered.

Journal Article↗

Antivirogram or phenosense: a comparison of their reproducibility and an analysis of their correlation.

The Antivirogram and PhenoSense assays are widely used phenotypic tests for HIV drug resistance. There are limited data on the reproducibility of each assay, and little is known about the correlation between the two. Using data from the Stanford HIV drug resistance database, we performed a comprehensive analysis of the reproducibility of each assay, and calculated the correlation and concordance of the two assays using both general IC50 fold change cutoff values and drug-specific cutoff values. Although the within-assay correlations were high (rank correlation coefficients r=0.94 and r=0.95 for the Antivirogram and PhenoSense assays, respectively), the between-assay correlation was considerably lower (r=0.36). Using drug-specific cutoff values for viruses classified as resistant by the Antivirogram or PhenoSense assays, respectively, only 71.4% [95% confidence intervals (95% CI): 58.7-82.1%] and 57.0% (95% CI: 45.3-68.1%) of the samples were classified as resistant using the other assay. The poor agreement between the assays was primarily due to the extremely poor correlation between these assays for samples with low resistance values (r=0.02 and r=0.61 for samples with the Antivirogram measurements lower or higher than 2.0, respectively). Since the cutoff values for both assays are relatively low, our analysis suggests that one should be very careful when interpreting measurements that are near the cutoff values for drug resistance.

Anti-HIV Agents↗

Reproducibility of low and high concentration data in reversed-phase liquid chromatography. II. Overloaded band profiles on Chromolith-C18.

Single-component adsorption isotherm data were acquired by frontal analysis (FA) for six low molecular weight compounds (phenol, aniline, caffeine, o-toluidine, p-toluidine and propylbenzoate) on one Chromolith-C18 column (#30, Merck, Darmstadt, Germany), using different methanol:water solutions (composition between 60/40 and 15/85 v/v, depending on the solute) as the mobile phase. These data were modeled for best agreement between the experimental data points and the adsorption isotherm model. The adsorption-energy distributions were also derived and used for the selection of the best isotherm model. Widely different models were obtained for these six compounds, four being convex upward (i.e., Langmuirian) and two having at least one inflection point. Overloaded band profiles corresponding to two different sample sizes (a low and a high loading factor) were recorded on six monolithic columns (#30-35) belonging to the same manufactured lot. These experimental band profiles were compared to the profiles calculated from the isotherm measured by FA on the first column, using the equilibrium-dispersive (ED) model of chromatography. For four of the six columns (#30, #32, #33, and #35), the reproducibility was better than 5 and 2.5% for the low and the high concentration profiles, respectively. On the other two columns (#31 and #34), the bands showed significant and systematic retention time shifts for all six compounds (with nearly identical band shapes), the relative adsorption being between 6 and 15% stronger on column #31 or between 2 and 7% lower on column #34. These differences seem to be correlated with the differences in the total porosities of these columns, which differ by 3% from columns #31 to #34, the higher porosity column giving the stronger adsorption.

Adsorption↗

Comparative analysis of amplified and nonamplified RNA for hybridization in cDNA microarray.

Limiting amounts of RNA is a major issue in cDNA microarray, especially when one is dealing with fresh tissue samples. Here we describe a protocol based on template switch and T7 amplification that led to efficient and linear amplification of 1300x. Using a glass-array containing 368 genes printed in three or six replicas covering a wide range of expression levels and ratios, we determined quality and reproducibility of the data obtained from one nonamplified and two independently amplified RNAs (aRNA) derived from normal and tumor samples using replicas with dye exchange (dye-swap measurements). Overall, signal-to-noise ratio improved when we used aRNA (1.45-fold for channel 1 and 2.02-fold for channel 2), increasing by 6% the number of spots with meaningful data. Measurements arising from independent aRNA samples showed strong correlation among themselves (r(2)=0.962) and with those from the nonamplified sample (r(2)=0.975), indicating the reproducibility and fidelity of the amplification procedure. Measurement differences, i.e, spots with poor correlation between amplified and nonamplified measurements, did not show association with gene sequence, expression intensity, or expression ratio and can, therefore, be compensated with replication. In conclusion, aRNA can be used routinely in cDNA microarray analysis, leading to improved quality of data with high fidelity and reproducibility.

Gene Expression Profiling↗

Quantitative in vitro assessment of N-alkyl sulphate-induced cytotoxicity in human keratinocytes (HaCaT). Comparison with in vivo human irritation tests.

A spontaneously immortalized human keratinocyte line, HaCaT, was used as an in vitro model to predict the cutaneous irritation of anionic surfactants. For this purpose, a number of sodium salts of N-alkyl sulphates with hydrocarbon chain lengths varying between C8 and C16 were studied for possible cytotoxic effects. The endpoints used to assess toxicity were uptake of the vital dye neutral red (NR) and cell morphology criteria 24 h after dosing. A linear proportionality between keratinocyte number and NR uptake was established. All tested surfactants had cytotoxic effects as demonstrated by a decreased NR uptake, which showed a clear dose-response relationship. Concentrations resulting in 50% inhibition of NR uptake (IC-50) ranged from 0.15 mmol (sodium lauryl sulphate, C12) to 1.23 mmol (sodium octyl sulphate, C8). The in vitro cytotoxicity data were highly reproducible when the test was repeated after several weeks. The cytotoxicity data from these assays were compared with the irritant responses (as evaluated by measurement of erythema and transepidermal water loss) obtained after 24 h application of the same compounds (300 microliters of 20 mmol aqueous solution) to the volar forearm of human volunteers. There were significant linear correlations between the IC-50 values and both barrier damage (transepidermal water loss) and erythema (as evaluated by skin colour reflectance measurements). For the test substances, however, the sensitivity of the in vitro system was between 10 and 100 times higher than that observed in human skin in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Line↗

Design, validation, and application of a seven-strain Staphylococcus aureus PCR product microarray for comparative genomics.

Bacterial comparative genomics has been revolutionized by microarrays, but the power of any microarray is dependent on the number and diversity of gene reporters it contains. Staphylococcus aureus is an important human pathogen causing a wide range of invasive and toxin-mediated diseases, and more than 20% of the genome of any isolate consists of variable genes. Seven whole-genome sequences of S. aureus are available, and we exploited this rare opportunity to design, build, and validate a comprehensive, nonredundant PCR product microarray carrying reporters that represent every predicted open reading frame (3,623 probes). Such a comprehensive microarray necessitated a novel design strategy. Validation with the seven sequenced strains showed correct identification of 93.9% of genes present or absent/divergent but was dependent on the method of analysis chosen. Microarray data were highly reproducible, reducing the need for many replicate slides. Interpretation of microarray data was enhanced by focusing on the major areas of variation--the presence or absence of mobile genetic elements (MGEs). We compiled "composite genomes" of every individual MGE and visualized their distribution. This allowed the sensitive discrimination of related isolates, including the first clear description of how isolates of the same clone of epidemic methicillin-resistant S. aureus differ substantially in their carriage of MGEs. These MGEs carry virulence and resistance genes, suggesting differences in pathogenic potential. The novel methods of design and interpretation of data generated from this microarray will enable further studies of S. aureus evolution, epidemiology, and pathogenesis.

Bacterial Proteins↗

IUPAC gas chromatographic method for determination of fatty acid composition: collaborative study.

An international collaborative study of IUPAC methods II.D.19 and II.D.25 for preparation and GLC analysis of fatty acid methyl esters was begun in 1976. The IUPAC methodology, applicable to animal and vegetable oils and fats and fatty acids from all sources, contains special instructions for preparation and analysis of methyl esters of fatty acids containing 4 or more carbon atoms (analysis of milk fat). Twenty-three collaborators participated in the analysis of 5 known mixtures, 4 vegetable oils, 1 fish oil, and 2 butterfats. Several blind duplicate samples were included. The experimental data were subjected to statistical analysis to examine intra- and interlaboratory variation. Reproducibility and accuracy data for the higher fatty acid (14:0-22:1) mixtures and fish and vegetable oils were satisfactory and were in good agreement with results from an AOCS Smalley Committee check sample program involving analysis of the same samples. Typical coefficients of variation (%) at various concentrations were 15 (2% level), 8.5 (5% level), 7 (10% level), and 3 (50% level). Low recoveries and poor reproducibility were characteristics of results obtained for butyric acid in the butterfat and related known mixtures. A coefficient of variation of about 19% was found for analysis of butyric acid in butterfat, vs coefficients of variation in the range of 4-13% for similar levels of other components in butterfat and other samples. The IUPAC methodology for GLC analysis of fats and oils other than milk fats has been adopted by the AOAC as official first action to replace the current GLC method, 28.063-28.067.

Animals↗

Reproducibility of BOLD-based functional MRI obtained at 4 T.

The reproducibility of activation patterns in the whole brain obtained by functional magnetic resonance imaging (fMRI) experiments at 4 Tesla was studied with a simple finger-opposition task. Six subjects performed three runs in one session, and each run was analyzed separately with the t-test as a univariate method and Fisher's linear discriminant analysis as a multivariate method. Detrending with a first- and third-order polynomial as well as logarithmic transformation as preprocessing steps for the t-test were tested for their impact on reproducibility. Reproducibility across the whole brain was studied by using scatter plots of statistical values and calculating the correlation coefficient between pairs of activation maps. In order to compare reproducibility of "activated" voxels across runs, subjects and models, 2% of all voxels in the brain with the highest statistical values were classified as activated. The analysis of reproducible activated voxels was performed for the whole brain and within regions of interest. We found considerable variability in reproducibility across subjects, regions of interest, and analysis methods. The t-test on the linear detrended data yielded better reproducibility than Fisher's linear discriminant analysis, and therefore seems to be a robust although conservative method. Preliminary data indicate that these modeling results may be reversed by preprocessing to reduce respiratory and cardiac physiological noise effects. The reproducibility of both the position and number of activated voxels in the sensorimotor cortex was highest, while that of the supplementary motor area was much lower, with reproducibility of the cerebellum falling in between the other two areas.

Adult↗

Three-parameter lognormal distribution ubiquitously found in cDNA microarray data and its application to parametric data treatment.

BACKGROUND: To cancel experimental variations, microarray data must be normalized prior to analysis. Where an appropriate model for statistical data distribution is available, a parametric method can normalize a group of data sets that have common distributions. Although such models have been proposed for microarray data, they have not always fit the distribution of real data and thus have been inappropriate for normalization. Consequently, microarray data in most cases have been normalized with non-parametric methods that adjust data in a pair-wise manner. However, data analysis and the integration of resultant knowledge among experiments have been difficult, since such normalization concepts lack a universal standard. RESULTS: A three-parameter lognormal distribution model was tested on over 300 sets of microarray data. The model treats the hybridization background, which is difficult to identify from images of hybridization, as one of the parameters. A rigorous coincidence of the model to data sets was found, proving the model's appropriateness for microarray data. In fact, a closer fitting to Northern analysis was obtained. The model showed inconsistency only at very strong or weak data intensities. Measurement of z-scores as well as calculated ratios was reproducible only among data in the model-consistent intensity range; also, the ratios were independent of signal intensity at the corresponding range. CONCLUSION: The model could provide a universal standard for data, simplifying data analysis and knowledge integration. It was deduced that the ranges of inconsistency were caused by experimental errors or additive noise in the data; therefore, excluding the data corresponding to those marginal ranges will prevent misleading analytical conclusions.

Blotting, Northern↗

Cardiac sodium channel Markov model with temperature dependence and recovery from inactivation.

A Markov model of the cardiac sodium channel is presented. The model is similar to the CA1 hippocampal neuron sodium channel model developed by Kuo and Bean (1994. Neuron. 12:819-829) with the following modifications: 1) an additional open state is added; 2) open-inactivated transitions are made voltage-dependent; and 3) channel rate constants are exponential functions of enthalpy, entropy, and voltage and have explicit temperature dependence. Model parameters are determined using a simulated annealing algorithm to minimize the error between model responses and various experimental data sets. The model reproduces a wide range of experimental data including ionic currents, gating currents, tail currents, steady-state inactivation, recovery from inactivation, and open time distributions over a temperature range of 10 degrees C to 25 degrees C. The model also predicts measures of single channel activity such as first latency, probability of a null sweep, and probability of reopening.

Algorithms↗

Subsite mapping of enzymes: collecting and processing experimental data--a case study of an amylase-malto-oligosaccharide system.

Two research groups have independently developed the theory and experimental methodology for quantitatively assessing substrate monomer-subsite binding-energies for depolymerases. When the two approaches are applied to the same enzyme-substrate system they yield surprisingly divergent results. This paper outlines the application of the two approaches to an amylase-maltooligosaccharide system and points out the more important areas of disagreement. We show that by proper data-management, the conflicts between the tow laboratories are basically resolved. The complexities of the subsite model demand extensive data-gathering and exacting data-processing and verification that the computed model-parameters can faithfully reproduce the experimental data.

Amylases↗

Continuous ice core melter system with discrete sampling for major ion, trace element and stable isotope analyses.

We present a novel ice/firn core melter system that uses fraction collectors to collect discrete, high-resolution (<1 cm/sample possible), continuous, coregistered meltwater samples for analysis of eight major ions by ion chromatography (IC), >32 trace elements by inductively coupled plasma sectorfield mass spectrometry (ICP-SMS), and stable oxygen and hydrogen isotopes by isotope ratio mass spectrometry (IRMS). The new continuous melting with discrete sampling (CMDS) system preserves an archive of each sample, reduces the problem of incomplete particle dissolution in ICP-SMS samples, and provides more precise trace element data than previous ice melter models by using longer ICP-SMS scan times and washing the instrument between samples. CMDS detection limits are similar to or lower than those published for ice melter systems coupled directly to analytical instruments and are suitable for analyses of polar and mid-low-latitude ice cores. Analysis of total calcium and sulfur by ICP-SMS and calcium ion, sulfate, and methanesulfonate by IC from the Mt. Logan Prospector-Russell Col ice core confirms data accuracy and coregistration of the split fractions from each sample. The reproducibility of all data acquired by the CMDS system is confirmed by replicate analyses of parallel sections of the GISP2 D ice core.

Chromatography, Ion Exchange↗