Validation of a food frequency questionnaire for women with osteoporosis

The importance of nutrient intakes in osteoporosis prevention in treatment is widely recognized. The objective of the present study was to develop and validate a FFQ for women with osteoporosis. The questionnaire was composed of 60 items, separated into 10 groups. The relative validation was accomplished through comparison of the 3-Day Food Record (3DR) with the FFQ. The 3DR was applied to 30 elderly women with confirmed osteoporosis, and after 45 days the FFQ was administrated. Statistical analysis comprised the Kolmogorov-Smirnov, Student T test and Pearson correlation coefficient. The agreement between two methods was evaluated by the frequency of similar classification into quartiles, and by the Bland-Altman method. No significant differences between methods were observed for the mean evaluated nutrients, except for carbohydrate and magnesium. Pearson correlation coefficients were positive and statistically significant for all nutrients. The overall proportion of subjects classified in the same quartile by the two methods was on average 50.01% and in the opposite quartile 0.47%. For calcium intake, only 3% of subjects were classified in opposite extreme quartiles by the two methods. The Bland-Altman analysis demonstrated that the differences obtained by the two methods in each subject were well distributed around the mean of the difference, and the disagreement increases as the mean intake increases. These results indicates that the FFQ for elderly women with osteoporosis presented here is highly acceptable and is an accurate method that can be used in large-scale or clinical studies for evaluation of nutrient intakes in a similar population.

Validation of a novel ELISA for measurement of electronegative low-density lipoprotein

Background: Oxidative modification of low-density lipoprotein (LDL) plays a key role in the pathogenesis of atherosclerosis. LDL(-) is present in blood plasma of healthy subjects and at higher concentrations in diseases with high cardiovascular risk, such as familial hypercholesterolemia or diabetes. Methods: We developed and validated a sandwich ELISA for LDL(-) in human plasma using two monoclonal antibodies against LDL(-) that do not bind to native LDL, extensively copper-oxidized LDL or malondialdehyde-modified LDL. The characteristics of assay performance, such as limits of detection and quantification, accuracy, inter- and intra-assay precision were evaluated. The linearity, interferences and stability tests were also performed. Results: The calibration range of the assay is 0.625-20.0 mU/L at 1: 2000 sample dilution. ELISA validation showed intra- and inter- assay precision and recovery within the required limits for immunoassays. The limits of detection and quantification were 0.423 mU/L and 0.517 mU/L LDL(-), respectively. The intra- and inter- assay coefficient of variation ranged from 9.5% to 11.5% and from 11.3% to 18.9%, respectively. Recovery of LDL(-) ranged from 92.8% to 105.1%. Conclusions: This ELISA represents a very practical tool for measuring LDL(-) in human blood for widespread research and clinical sample use. Clin Chem Lab Med 2008; 46: 1769-75.

Validation of an ELISA method for the serological diagnosis of canine brucellosis due to Brucella canis

In the present study, the validation of an enzyme-linked immunosorbent assay (ELISA) for serodiagnosis of canine brucellosis is described. Two different antigenic extracts, obtained by heat or ultrasonic homogenization of microbial antigens from a wild isolate of Brucella canis bacteria, were compared by ELISA and Western blot (WB). A total of 145 canine sera were used to define sensitivity, specificity and accuracy of the ELISA as follows: (1) sera from 34 animals with natural B. canis infection, confirmed by blood culture and PCR, as well as 51 sera samples from healthy dogs with negative results by the agar-gel immunodiffusion (ACID) test for canine brucellosis, were used as the control panel for B. cants infection; and (2) to scrutinize the possibility of cross reactions with other common dog infections in the same geographical area in Brazil, 60 sera samples from dogs harboring known infections by Leptospira sp., Ehrlichia canis, canine distemper virus (CDV), Neospora caninum, Babesia canis and Leishmania chagasi (10 in each group) were included in the study. The ELISA using heat soluble bacterial extract (HE-antigen) as antigen showed the best values of sensitivity (91.18%), specificity (100%) and accuracy (96.47%). In the WB analyses, the HE-antigen showed no cross-reactivity with sera from dogs with different infections, while the B. canis sonicate had various protein bands identified by those sera. The performance of the ELISA standardized with the heat soluble B. canis antigen indicates that this assay can be used as a reliable and practical method to confirm infection by this microorganism, as well as a tool for seroepidemiological studies. (C) 2010 Elsevier Ltd. All rights reserved.

Phylogenetic Validation of the Genera Angomonas and Strigomonas of Trypanosomatids Harboring Bacterial Endosymbionts with the Description of New Species of Trypanosomatids and of Proteobacterial Symbionts

We comparatively examined the nutritional, molecular and optical and electron microscopical characteristics of reference species and new isolates of trypanosomatids harboring bacterial endosymbionts. Sequencing of the V7V8 region of the small subunit of the ribosomal RNA (SSU rRNA) gene distinguished six major genotypes among the 13 isolates examined. The entire sequences of the SSU rRNA and glycosomal glyceraldehyde phosphate dehydrogenase (gGAPDH) genes were obtained for phylogenetic analyses. In the resulting phylogenetic trees, the symbiont-harboring species clustered as a major clade comprising two subclades that corresponded to the proposed genera Angomonas and Strigomonas. The genus Angomonas comprised 10 flagellates including former Crithidia deanei and C. desouzai plus a new species. The genus Strigomonas included former Crithidia oncopelti and Blastocrithidia cuiicis plus a new species. Sequences from the internal transcribed spacer of ribosomal DNA (ITS rDNA) and size polymorphism of kinetoplast DNA (kDNA) minicircles revealed considerable genetic heterogeneity within the genera Angomonas and Strigomonas. Phylogenetic analyses based on 16S rDNA and ITS rDNA sequences demonstrated that all of the endosymbionts belonged to the Betaproteobacteria and revealed three new species. The congruence of the phylogenetic trees of trypanosomatids and their symbionts support a co-divergent host-symbiont evolutionary history. (C) 2011 Elsevier GmbH. All rights reserved.

Sequence-specific reconstruction from fragmentary databases using seed sequences: implementation and validation on SAGE, proteome and generic sequencing data

Motivation: DNA assembly programs classically perform an all-against-all comparison of reads to identify overlaps, followed by a multiple sequence alignment and generation of a consensus sequence. If the aim is to assemble a particular segment, instead of a whole genome or transcriptome, a target-specific assembly is a more sensible approach. GenSeed is a Perl program that implements a seed-driven recursive assembly consisting of cycles comprising a similarity search, read selection and assembly. The iterative process results in a progressive extension of the original seed sequence. GenSeed was tested and validated on many applications, including the reconstruction of nuclear genes or segments, full-length transcripts, and extrachromosomal genomes. The robustness of the method was confirmed through the use of a variety of DNA and protein seeds, including short sequences derived from SAGE and proteome projects.

Formal validation of automated policy refinement in the management of network security systems

Policy hierarchies and automated policy refinement are powerful approaches to simplify administration of security services in complex network environments. A crucial issue for the practical use of these approaches is to ensure the validity of the policy hierarchy, i.e. since the policy sets for the lower levels are automatically derived from the abstract policies (defined by the modeller), we must be sure that the derived policies uphold the high-level ones. This paper builds upon previous work on Model-based Management, particularly on the Diagram of Abstract Subsystems approach, and goes further to propose a formal validation approach for the policy hierarchies yielded by the automated policy refinement process. We establish general validation conditions for a multi-layered policy model, i.e. necessary and sufficient conditions that a policy hierarchy must satisfy so that the lower-level policy sets are valid refinements of the higher-level policies according to the criteria of consistency and completeness. Relying upon the validation conditions and upon axioms about the model representativeness, two theorems are proved to ensure compliance between the resulting system behaviour and the abstract policies that are modelled.

Development and Validation of a Mice liar Electrokinetic Chromatography Method for Quantitative Determination of Butenolides in Piper malacophyllum (C. Presl) C. DC.

Introduction - A large number of natural and synthetic compounds having butenolides as a core unit have been described and many of them display a wide range of biological activities. Butenolides from P. malacophyllum have presented potential antifungal activities but no specific, fast, and precise method has been developed for their determination. Objective - To develop a methodology based on micellar electrokinetic chromatography to determine butenolides in Piper species. Methodology - The extracts were analysed in an uncoated fused-silica capillaries and for the micellar system 20 mmol/L SDS, 20% (v/v) acetonitrile (ACN) and 10 mmol/L STB aqueous buffer at pH 9.2 were used. The method was validated for precision, linearity, limit of detection (LOD) and limit of quantitation (LOQ) and the standard deviations were determined from the standard errors estimated by the regression line. Results - A micellar electrokinetic chromatography (MEKC) method for determination of butenolides in extracts gave full resolution for 1 and 2. The analytical curve in the range 10.0-50.0 mu g/mL (r(2) = 0.999) provided LOD and LOQ for 1 and 2 of 2.1/6.3 and 1.1/3.5 mu g/mL, respectively. The RSD for migration times were 0.12 and 1.0% for peak area ratios with 100.0 +/- 1.4% of recovery. Conclusions - A novel high-performance MEKC method developed for the analysis of butenolides 1 and 2 in leaf extracts of P. malacophyllum allowed their quantitative determined within an analysis time shorter than 5 min and the results indicated CE to be a feasible analytical technique for the quantitative determination of butenolides in Piper extracts. Copyright (C) 2010 John Wiley & Sons, Ltd.

Multivariant optimization, validation, and application of capillary electrophoresis for simultaneous determination of polyphenols and phenolic acids in Brazilian wines

A method for the simultaneous determination of the stilbene resveratrol, four phenolic acids (syringic, coumaric, caffeic, and gallic acids), and five flavonoids (catechin, rutin, kaempferol, myricetin, and quercetin) in wine by CE was developed and validated. The CE electrolyte composition and instrumental conditions were optimized using 2(7-3) factorial design and response surface analysis, showing sodium tetraborate, MeOH, and their interaction as the most influential variables. The optimal electrophoretic conditions, minimizing the chromatographic resolution statistic values, consisted of 17 mmol/L sodium tetraborate with 20% methanol as electrolyte, constant voltage of 25 kV, hydrodynamic injection at 50 mbar for 3 s, and temperature of 25 degrees C. The R(2) values for linearity varied from 0.994 to 0.999; LOD and LOQ were 0.1 to 0.3 mg/L and 0.4 to 0.8 mg/L, respectively. The RSDs for migration time and peak area obtained from ten consecutive injections were less than 2% and recoveries varied from 97 to 102%. The method was applied to 23 samples of inexpensive Brazilian wines, showing wide compositional variation.