Inter-laboratory validation of bioaccessibility testing for metals.

PubMed

Henderson, Rayetta G; Verougstraete, Violaine; Anderson, Kim; Arbildua, José J; Brock, Thomas O; Brouwers, Tony; Cappellini, Danielle; Delbeke, Katrien; Herting, Gunilla; Hixon, Greg; Odnevall Wallinder, Inger; Rodriguez, Patricio H; Van Assche, Frank; Wilrich, Peter; Oller, Adriana R

2014-10-01

Bioelution assays are fast, simple alternatives to in vivo testing. In this study, the intra- and inter-laboratory variability in bioaccessibility data generated by bioelution tests were evaluated in synthetic fluids relevant to oral, inhalation, and dermal exposure. Using one defined protocol, five laboratories measured metal release from cobalt oxide, cobalt powder, copper concentrate, Inconel alloy, leaded brass alloy, and nickel sulfate hexahydrate. Standard deviations of repeatability (sr) and reproducibility (sR) were used to evaluate the intra- and inter-laboratory variability, respectively. Examination of the sR:sr ratios demonstrated that, while gastric and lysosomal fluids had reasonably good reproducibility, other fluids did not show as good concordance between laboratories. Relative standard deviation (RSD) analysis showed more favorable reproducibility outcomes for some data sets; overall results varied more between- than within-laboratories. RSD analysis of sr showed good within-laboratory variability for all conditions except some metals in interstitial fluid. In general, these findings indicate that absolute bioaccessibility results in some biological fluids may vary between different laboratories. However, for most applications, measures of relative bioaccessibility are needed, diminishing the requirement for high inter-laboratory reproducibility in absolute metal releases. The inter-laboratory exercise suggests that the degrees of freedom within the protocol need to be addressed. Copyright © 2014 Elsevier Inc. All rights reserved.

Interlaboratory comparison of chemical analysis of uranium mononitride

NASA Technical Reports Server (NTRS)

Merkle, E. J.; Davis, W. F.; Halloran, J. T.; Graab, J. W.

1974-01-01

Analytical methods were established in which the critical variables were controlled, with the result that acceptable interlaboratory agreement was demonstrated for the chemical analysis of uranium mononitride. This was accomplished by using equipment readily available to laboratories performing metallurgical analyses. Agreement among three laboratories was shown to be very good for uranium and nitrogen. Interlaboratory precision of + or - 0.04 percent was achieved for both of these elements. Oxygen was determined to + or - 15 parts per million (ppm) at the 170-ppm level. The carbon determination gave an interlaboratory precision of + or - 46 ppm at the 320-ppm level.

Interlaboratory comparison of real-time pcr protocols for quantification of general fecal indicator bacteria

USGS Publications Warehouse

Shanks, O.C.; Sivaganesan, M.; Peed, L.; Kelty, C.A.; Blackwood, A.D.; Greene, M.R.; Noble, R.T.; Bushon, R.N.; Stelzer, E.A.; Kinzelman, J.; Anan'Eva, T.; Sinigalliano, C.; Wanless, D.; Griffith, J.; Cao, Y.; Weisberg, S.; Harwood, V.J.; Staley, C.; Oshima, K.H.; Varma, M.; Haugland, R.A.

2012-01-01

The application of quantitative real-time PCR (qPCR) technologies for the rapid identification of fecal bacteria in environmental waters is being considered for use as a national water quality metric in the United States. The transition from research tool to a standardized protocol requires information on the reproducibility and sources of variation associated with qPCR methodology across laboratories. This study examines interlaboratory variability in the measurement of enterococci and Bacteroidales concentrations from standardized, spiked, and environmental sources of DNA using the Entero1a and GenBac3 qPCR methods, respectively. Comparisons are based on data generated from eight different research facilities. Special attention was placed on the influence of the DNA isolation step and effect of simplex and multiplex amplification approaches on interlaboratory variability. Results suggest that a crude lysate is sufficient for DNA isolation unless environmental samples contain substances that can inhibit qPCR amplification. No appreciable difference was observed between simplex and multiplex amplification approaches. Overall, interlaboratory variability levels remained low (<10% coefficient of variation) regardless of qPCR protocol. ?? 2011 American Chemical Society.

Fuzzy-logic based strategy for validation of multiplex methods: example with qualitative GMO assays.

PubMed

Bellocchi, Gianni; Bertholet, Vincent; Hamels, Sandrine; Moens, W; Remacle, José; Van den Eede, Guy

2010-02-01

This paper illustrates the advantages that a fuzzy-based aggregation method could bring into the validation of a multiplex method for GMO detection (DualChip GMO kit, Eppendorf). Guidelines for validation of chemical, bio-chemical, pharmaceutical and genetic methods have been developed and ad hoc validation statistics are available and routinely used, for in-house and inter-laboratory testing, and decision-making. Fuzzy logic allows summarising the information obtained by independent validation statistics into one synthetic indicator of overall method performance. The microarray technology, introduced for simultaneous identification of multiple GMOs, poses specific validation issues (patterns of performance for a variety of GMOs at different concentrations). A fuzzy-based indicator for overall evaluation is illustrated in this paper, and applied to validation data for different genetically modified elements. Remarks were drawn on the analytical results. The fuzzy-logic based rules were shown to be applicable to improve interpretation of results and facilitate overall evaluation of the multiplex method.

Interlaboratory round robin study on axial tensile properties of SiC-SiC CMC tubular test specimens [Interlaboratory round robin study on axial tensile properties of SiC/SiC tubes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Singh, Gyanender P.; Gonczy, Steve T.; Deck, Christian P.

An interlaboratory round robin study was conducted on the tensile strength of SiC–SiC ceramic matrix composite (CMC) tubular test specimens at room temperature with the objective of expanding the database of mechanical properties of nuclear grade SiC–SiC and establishing the precision and bias statement for standard test method ASTM C1773. The mechanical properties statistics from the round robin study and the precision statistics and precision statement are presented herein. The data show reasonable consistency across the laboratories, indicating that the current C1773–13 ASTM standard is adequate for testing ceramic fiber reinforced ceramic matrix composite tubular test specimen. Furthermore, it wasmore » found that the distribution of ultimate tensile strength data was best described with a two–parameter Weibull distribution, while a lognormal distribution provided a good description of the distribution of proportional limit stress data.« less

Interlaboratory round robin study on axial tensile properties of SiC-SiC CMC tubular test specimens [Interlaboratory round robin study on axial tensile properties of SiC/SiC tubes

DOE PAGES

Singh, Gyanender P.; Gonczy, Steve T.; Deck, Christian P.; ...

2018-04-19

An interlaboratory round robin study was conducted on the tensile strength of SiC–SiC ceramic matrix composite (CMC) tubular test specimens at room temperature with the objective of expanding the database of mechanical properties of nuclear grade SiC–SiC and establishing the precision and bias statement for standard test method ASTM C1773. The mechanical properties statistics from the round robin study and the precision statistics and precision statement are presented herein. The data show reasonable consistency across the laboratories, indicating that the current C1773–13 ASTM standard is adequate for testing ceramic fiber reinforced ceramic matrix composite tubular test specimen. Furthermore, it wasmore » found that the distribution of ultimate tensile strength data was best described with a two–parameter Weibull distribution, while a lognormal distribution provided a good description of the distribution of proportional limit stress data.« less

Determination of perfluorinated alkyl acid concentrations in human serum and milk standard reference materials.

PubMed

Keller, Jennifer M; Calafat, Antonia M; Kato, Kayoko; Ellefson, Mark E; Reagen, William K; Strynar, Mark; O'Connell, Steven; Butt, Craig M; Mabury, Scott A; Small, Jeff; Muir, Derek C G; Leigh, Stefan D; Schantz, Michele M

2010-05-01

Standard Reference Materials (SRMs) are certified reference materials produced by the National Institute of Standards and Technology that are homogeneous materials well characterized with values for specified properties, such as environmental contaminant concentrations. They can be used to validate measurement methods and are critical in improving data quality. Disagreements in perfluorinated alkyl acid (PFAA) concentrations measured in environmental matrices during past interlaboratory comparisons emphasized the need for SRMs with values assigned for PFAAs. We performed a new interlaboratory comparison among six laboratories and provided, for the first time, value assignment of PFAAs in SRMs. Concentrations for perfluorooctane sulfonate (PFOS), perfluorooctanoate (PFOA), and other PFAAs in two human serum and two human milk SRMs are reported. PFAA concentration measurements agreed for serum SRM 1957 using different analytical methods in six laboratories and for milk SRM 1954 in three laboratories. The interlaboratory relative standard deviation for PFOS in SRM 1957 was 7%, which is an improvement over past interlaboratory studies. Matrix interferences are discussed, as well as temporal trends and the percentage of branched vs. linear isomers. The concentrations in these SRMs are similar to the present-day average concentrations measured in human serum and milk, resulting in representative and useful control materials for PFAA human monitoring studies.

Results of a European interlaboratory comparison on CO2 sorption on activated carbon and coals

NASA Astrophysics Data System (ADS)

Gensterblum, Yves; Busch, Andreas; Krooss, Bernhard; de Weireld, Guy; Billemont, Pierre; van Hemert, Patrick; Wolf, Karl-Heinz

2013-04-01

For the assessment of CO2 storage in coal seams or enhanced coalbed methane production (ECBM), the sorption properties of natural coals are important parameters. Since more and more laboratories worldwide are concerned with measurements of gas sorption on coal it is indispensable to establish quality standards for such experiments. The first two interlaboratory studies on CO2 sorption on coal (Goodman et al. 2004, 2007) revealed a poor agreement of sorption isotherms among the participating laboratories, particularly in the high-pressure range. During the MOVECBM (http://www.movecbm.eu/) project funded by the European Commission (6th framework), an interlaboratory comparison of CO2 sorption on selected coals and activated carbon was initiated. Measurements were performed on dry samples at 45° C using the manometric and the gravimetric method. up to a final pressure of 15 MPa. The first set of high-pressure sorption measurements was performed on a Filtrasorb 400 activated carbon sample in order to minimise heterogeneity effects and to optimize the experimental procedures for the individual (manometric or gravimetric) methods (Gensterblum et al. 2009). Since comparability for the activated carbon was excellent, the measurements were continued using natural coals of various rank (anthracite, bituminous coal and lignite) to study the influence of heterogeneities and varying starting conditions on the CO2 sorption properties (Gensterblum et al. 2010). Compared to the poor reproducibility observed in previous interlaboratory studies (Goodman et al., 2004, 2007) this European study showed excellent agreement (<5 % deviation) among the participating laboratories with good repeatability. The sorption data and technical information on the different experimental setups have been used to investigate errors and potential pitfalls in the assessment of high-pressure CO2 sorption isotherms. References Gensterblum Y., P. van Hemert, P. Billemont, A. Busch, B.M. Krooss, G. de Weireld, D. Prinz , K.-H.A.A. Wolf, "European inter-laboratory comparison of high pressure CO2 sorption isotherms. II: natural coals" IJCG, 2010, 84, 115-124 Gensterblum Y., P. van Hemert, P. Billemont, A. Busch, D. Charriére, D. Li, B.M. Krooss, G. de Weireld, D. Prinz , K.-H.A.A. Wolf, "European inter-laboratory comparison of high pressure CO2 sorption isotherms. I: Activated carbon" Carbon 47 ( 2009 ) 2958 -2969 Goodman, A.L., Busch, A., Duffy, G., Fitzgerald, J.E., Gasem, K.A.M., Gensterblum, Y., Krooss, B.M., Levy, J., Ozdemir, E., Pan, Z., Robinson, Jr., R.L., Schroeder, K., Sudibandriyo, M., White, C. (2004). An Inter-laboratory Comparison of CO2 Isotherms Measured on Argonne Premium Coal Samples. Energy and Fuels 18, 1175-1182. Goodman, A.L., Busch, A., Day, S., Duffy, G.J., Fitzgerald, J.E., Gasem, K.A.M., Gensterblum, Y., Hartman, C., Krooss, B.M., Pan, Z., Pratt, T., Robinson, Jr., R.L., Romanov, V., Sakurovs, R., Schroeder, K., Sudibandriyo, M., White, C.M. (2007) "Inter-laboratory Comparison II: CO2 Isotherms Measured on Moisture-Equilibrated Argonne Premium Coals at 55oC and 15 MPa", International Journal of Coal Geology 72, 153-164.

Determination of the moisture content of instant noodles: interlaboratory study.

PubMed

Hakoda, Akiko; Kasama, Hirotaka; Sakaida, Kenichi; Suzuki, Tadanao; Yasui, Akemi

2006-01-01

Determination of the moisture content of instant noodles, currently under discussion by the Codex Alimentarius Commission (CAC) requires 2 methods: one for fried noodles and the other for nonfried noodles. The method to determine the moisture content of fried noodles by drying at 105 degrees C for 2 h used in the Japanese Agricultural Standard (JAS) system of Japan can be applied to this purpose. In the present study, the JAS method for fried noodles was modified to be suitable for nonfried noodles by extending the drying time to 4 h. An interlaboratory study was conducted to evaluate interlaboratory performance statistics for these 2 methods. Ten participating laboratories each analyzed 5 test materials of fried and nonfried noodles as blind duplicates. After removal of outliers statistically, the repeatability (RSDr) and the reproducibility (RSD(R)) of these methods were 1.6-2.6 and 3.9-4.8% for fried noodles, and 0.3-1.5 and 1.3-2.9% for nonfried noodles, respectively.

Multielement trace determination in SiC powders: assessment of interlaboratory comparisons aimed at the validation and standardization of analytical procedures with direct solid sampling based on ETV ICP OES and DC arc OES.

PubMed

Matschat, Ralf; Hassler, Jürgen; Traub, Heike; Dette, Angelika

2005-12-01

The members of the committee NMP 264 "Chemical analysis of non-oxidic raw and basic materials" of the German Standards Institute (DIN) have organized two interlaboratory comparisons for multielement determination of trace elements in silicon carbide (SiC) powders via direct solid sampling methods. One of the interlaboratory comparisons was based on the application of inductively coupled plasma optical emission spectrometry with electrothermal vaporization (ETV ICP OES), and the other on the application of optical emission spectrometry with direct current arc (DC arc OES). The interlaboratory comparisons were organized and performed in the framework of the development of two standards related to "the determination of mass fractions of metallic impurities in powders and grain sizes of ceramic raw and basic materials" by both methods. SiC powders were used as typical examples of this category of material. The aim of the interlaboratory comparisons was to determine the repeatability and reproducibility of both analytical methods to be standardized. This was an important contribution to the practical applicability of both draft standards. Eight laboratories participated in the interlaboratory comparison with ETV ICP OES and nine in the interlaboratory comparison with DC arc OES. Ten analytes were investigated by ETV ICP OES and eleven by DC arc OES. Six different SiC powders were used for the calibration. The mass fractions of their relevant trace elements were determined after wet chemical digestion. All participants followed the analytical requirements described in the draft standards. In the calculation process, three of the calibration materials were used successively as analytical samples. This was managed in the following manner: the material that had just been used as the analytical sample was excluded from the calibration, so the five other materials were used to establish the calibration plot. The results from the interlaboratory comparisons were summarized and used to determine the repeatability and the reproducibility (expressed as standard deviations) of both methods. The calculation was carried out according to the related standard. The results are specified and discussed in this paper, as are the optimized analytical conditions determined and used by the authors of this paper. For both methods, the repeatability relative standard deviations were <25%, usually ~10%, and the reproducibility relative standard deviations were <35%, usually ~15%. These results were regarded as satifactory for both methods intended for rapid analysis of materials for which decomposition is difficult and time-consuming. Also described are some results from an interlaboratory comparison used to certify one of the materials that had been previously used for validation in both interlaboratory comparisons. Thirty laboratories (from eight countries) participated in this interlaboratory comparison for certification. As examples, accepted results are shown from laboratories that used ETV ICP OES or DC arc OES and had performed calibrations by using solutions or oxides, respectively. The certified mass fractions of the certified reference materials were also compared with the mass fractions determined in the interlaboratory comparisons performed within the framework of method standardization. Good agreement was found for most of the analytes.

Using an interlaboratory study to revise methods for conducting 10-d to 42-d water or sediment toxicity tests with Hyalella azteca

USGS Publications Warehouse

Ivey, Chris D.; Ingersoll, Christopher G.; Brumbaugh, William G.; Hammer, Edward J.; Mount, David R.; Hockett, J. Russell; Norberg-King, Teresa J.; Soucek, Dave; Taylor, Lisa

2016-01-01

Studies have been conducted to refine US Environmental Protection Agency, ASTM International, and Environment Canada standard methods for conducting 42-d reproduction tests with Hyalella azteca in water or in sediment. Modifications to the H. azteca method include better-defined ionic composition requirements for exposure water (i.e., >15 mg/L of chloride and >0.02 mg/L of bromide) and improved survival, growth, and reproduction with alternate diets provided as increased rations over time in water-only or whole-sediment toxicity tests. A total of 24 laboratories volunteered to participate in the present interlaboratory study evaluating the performance of H. azteca in 42-d studies in control sand or control sediment using the refined methods. Improved growth and reproduction of H. azteca was observed with 2 alternate diets of 1) ramped diatoms (Thalassiosira weissflogii) + ramped Tetramin or 2) yeast–cerophyll–trout chow (YCT) + ramped Tetramin, especially when compared with results from the traditional diet of 1.8 mg YCT/d. Laboratories were able to meet proposed test acceptability criteria and in most cases had lower variation in growth or reproduction compared with previous interlaboratory studies using the traditional YCT diet. Laboratory success in conducting 42-d H. azteca exposures benefited from adherence to several key requirements of the detailed testing, culturing, and handling methods. Results from the present interlaboratory study are being used to help revise standard methods for conducting 10-d to 42-d water or sediment toxicity exposures with H. azteca.

Results of interlaboratory comparison of fission track ages for 1992 fission track workshop

USGS Publications Warehouse

Miller, D.S.; Crowley, K.D.; Dokka, R.K.; Galbraith, R.F.; Kowallis, B.J.; Naeser, C.W.

1993-01-01

Two apatites and one sphene were made available to the fission track research community for analysis prior to the 1992 Fission Track Workshop held in Philadelphia, U.S.A., 13-17 July. Eighteen laboratories throughout the world received aliquots of apatite and sphene. To date, analyses by 33 different scientists have been representing 15 different laboratories. With respect to the previous two interlaboratory comparisons, there is a noticeable improvement in the accuracy of the age results (Naeser and Cebula, 1978; Naeser et al., 1981; Miller et al., 1985;Miller et al.1990). Ninety-four percent of the analysis used the external detector method (EDM) combined with the zeta technique while the remaining individuals used the population method (POP). Track length measurements (requested for the first time in the interlaboratory comparison studies) were in relatively good agreement. ?? 1993.

Statistical Analysis of a Round-Robin Measurement Survey of Two Candidate Materials for a Seebeck Coefficient Standard Reference Material

PubMed Central

Lu, Z. Q. J.; Lowhorn, N. D.; Wong-Ng, W.; Zhang, W.; Thomas, E. L.; Otani, M.; Green, M. L.; Tran, T. N.; Caylor, C.; Dilley, N. R.; Downey, A.; Edwards, B.; Elsner, N.; Ghamaty, S.; Hogan, T.; Jie, Q.; Li, Q.; Martin, J.; Nolas, G.; Obara, H.; Sharp, J.; Venkatasubramanian, R.; Willigan, R.; Yang, J.; Tritt, T.

2009-01-01

In an effort to develop a Standard Reference Material (SRM™) for Seebeck coefficient, we have conducted a round-robin measurement survey of two candidate materials—undoped Bi2Te3 and Constantan (55 % Cu and 45 % Ni alloy). Measurements were performed in two rounds by twelve laboratories involved in active thermoelectric research using a number of different commercial and custom-built measurement systems and techniques. In this paper we report the detailed statistical analyses on the interlaboratory measurement results and the statistical methodology for analysis of irregularly sampled measurement curves in the interlaboratory study setting. Based on these results, we have selected Bi2Te3 as the prototype standard material. Once available, this SRM will be useful for future interlaboratory data comparison and instrument calibrations. PMID:27504212

Interlaboratory studies and initiatives developing standards for proteomics

PubMed Central

Ivanov, Alexander R.; Colangelo, Christopher M.; Dufresne, Craig P.; Friedman, David B.; Lilley, Kathryn S.; Mechtler, Karl; Phinney, Brett S.; Rose, Kristie L.; Rudnick, Paul A.; Searle, Brian C.; Shaffer, Scott A.; Weintraub, Susan T.

2013-01-01

Proteomics is a rapidly transforming interdisciplinary field of research that embraces a diverse set of analytical approaches to tackle problems in fundamental and applied biology. This view-point article highlights the benefits of interlaboratory studies and standardization initiatives to enable investigators to address many of the challenges found in proteomics research. Among these initiatives, we discuss our efforts on a comprehensive performance standard for characterizing PTMs by MS that was recently developed by the Association of Biomolecular Resource Facilities (ABRF) Proteomics Standards Research Group (sPRG). PMID:23319436

Interlaboratory comparison of measurements of acid-volatile sulfide and simultaneously extracted nickel in spiked sediments

USGS Publications Warehouse

Brumbaugh, William G.; Hammerschmidt, Chad R.; Zanella, Luciana; Rogevich, Emily; Salata, Gregory; Bolek, Radoslaw

2011-01-01

An interlaboratory comparison of acid-volatile sulfide (AVS) and simultaneously extracted nickel (SEM_Ni) measurements of sediments was conducted among five independent laboratories. Relative standard deviations for the seven test samples ranged from 5.6 to 71% (mean = 25%) for AVS and from 5.5 to 15% (mean = 10%) for SEM_Ni. These results are in stark contrast to a recently published study that indicated AVS and SEM analyses were highly variable among laboratories.

Accuracy of trace element determinations in alternate fuels

NASA Technical Reports Server (NTRS)

Greenbauer-Seng, L. A.

1980-01-01

A review of the techniques used at Lewis Research Center (LeRC) in trace metals analysis is presented, including the results of Atomic Absorption Spectrometry and DC Arc Emission Spectrometry of blank levels and recovery experiments for several metals. The design of an Interlaboratory Study conducted by LeRC is presented. Several factors were investigated, including: laboratory, analytical technique, fuel type, concentration, and ashing additive. Conclusions drawn from the statistical analysis will help direct research efforts toward those areas most responsible for the poor interlaboratory analytical results.

Qualitative PCR method for Roundup Ready soybean: interlaboratory study.

PubMed

Kodama, Takashi; Kasahara, Masaki; Minegishi, Yasutaka; Futo, Satoshi; Sawada, Chihiro; Watai, Masatoshi; Akiyama, Hiroshi; Teshima, Reiko; Kurosawa, Yasunori; Furui, Satoshi; Hino, Akihiro; Kitta, Kazumi

2011-01-01

Quantitative and qualitative methods based on PCR have been developed for genetically modified organisms (GMO). Interlaboratory studies were previously conducted for GMO quantitative methods; in this study, an interlaboratory study was conducted for a qualitative method for a GM soybean, Roundup Ready soy (RR soy), with primer pairs designed for the quantitative method of RR soy studied previously. Fourteen laboratories in Japan participated. Each participant extracted DNA from 1.0 g each of the soy samples containing 0, 0.05, and 0.10% of RR soy, and performed PCR with primer pairs for an internal control gene (Le1) and RR soy followed by agarose gel electrophoresis. The PCR product amplified in this PCR system for Le1 was detected from all samples. The sensitivity, specificity, and false-negative and false-positive rates of the method were obtained from the results of RR soy detection. False-negative rates at the level of 0.05 and 0.10% of the RR soy samples were 6.0 and 2.3%, respectively, revealing that the LOD of the method was somewhat below 0.10%. The current study demonstrated that the qualitative method would be practical for monitoring the labeling system of GM soy in kernel lots.

Interlaboratory evaluation of trace element determination in workplace air filter samples by inductively coupled plasma mass spectrometry†‡

PubMed Central

Shulman, Stanley A.; Brisson, Michael J.; Howe, Alan M.

2015-01-01

Inductively coupled plasma mass spectrometry (ICP-MS) is becoming more widely used for trace elemental analysis in the occupational hygiene field, and consequently new ICP-MS international standard procedures have been promulgated by ASTM International and ISO. However, there is a dearth of interlaboratory performance data for this analytical methodology. In an effort to fill this data void, an interlaboratory evaluation of ICP-MS for determining trace elements in workplace air samples was conducted, towards fulfillment of method validation requirements for international voluntary consensus standard test methods. The study was performed in accordance with applicable statistical procedures for investigating interlaboratory precision. The evaluation was carried out using certified 37-mm diameter mixed-cellulose ester (MCE) filters that were fortified with 21 elements of concern in occupational hygiene. Elements were spiked at levels ranging from 0.025 to 10 μg filter−1, with three different filter loadings denoted “Low”, “Medium” and “High”. Participating laboratories were recruited from a pool of over fifty invitees; ultimately twenty laboratories from Europe, North America and Asia submitted results. Triplicates of each certified filter with elemental contents at three different levels, plus media blanks spiked with reagent, were conveyed to each volunteer laboratory. Each participant was also provided a copy of the test method which each participant was asked to follow; spiking levels were unknown to the participants. The laboratories were requested to prepare the filters by one of three sample preparation procedures, i.e., hotplate digestion, microwave digestion or hot block extraction, which were described in the test method. Participants were then asked to analyze aliquots of the prepared samples by ICP-MS, and to report their data in units of μg filter−1. Most interlaboratory precision estimates were acceptable for medium- and high-level spikes (RSD <25%), but generally yielded greater uncertainties than were anticipated at the outset of the study. PMID:22038017

Interlaboratory Collaborations in the Undergraduate Setting

ERIC Educational Resources Information Center

Megehee, Elise G.; Hyslop, Alison G.; Rosso, Richard J.

2005-01-01

A novel approach to cross-disciplinary and group learning, known as interlaboratory collaborations, was developed. The method mimics an industrial or research setting, fosters teamwork, and emphasizes the importance of good communication skills in the sciences.

Interlaboratory and between-specimen comparisons of diagnostic tests for leptospirosis in sheep and cattle.

PubMed

Fang, Fang; Collins-Emerson, Julie M; Heuer, Cord; Hill, Fraser I; Tisdall, David J; Wilson, Peter R; Benschop, Jackie

2014-11-01

A study was performed to investigate interlaboratory test agreement between a research and a commercial veterinary diagnostic laboratory on blood and urine samples, and to investigate test agreement between blood, urine, and kidney samples (research laboratory) for leptospirosis diagnosis. Samples were sourced from 399 sheep and 146 beef cattle from a local abattoir. Interlaboratory agreement for real-time quantitative polymerase chain reaction (qPCR) results on urine samples was almost perfect (kappa = 0.90), despite the use of different amplification targets (DNA gyrase subunit B gene vs. 16s ribosomal RNA gene), chemistries (SYTO9 vs. TaqMan probe), and pre-PCR processing. Interlaboratory agreement for microscopic agglutination test (MAT) positivity was almost perfect (kappa = 0.93) for Leptospira borgpetersenii serovar Hardjo subtype Hardjobovis (Hardjobovis) but moderate (kappa = 0.53) for Leptospira interrogans serovar Pomona (Pomona). Among animals that had different titers recorded, higher Hardjobovis and lower Pomona titers were reported by the commercial laboratory than by the research laboratory (P < 0.005). These interlaboratory comparisons can assist researchers and diagnosticians in interpreting the sometimes discrepant test results. Within the research laboratory, the comparison of qPCR results on urine and kidney showed almost perfect agreement (kappa = 0.84), suggesting that the qPCR on these 2 specimens can be used interchangeably. The agreement between MAT positivity and urine and kidney qPCR results was fair (kappa = 0.32 and kappa = 0.33, respectively). However, the prevalence ratio of urine and kidney qPCR positivity in Hardjobovis-seropositive versus Hardjobovis-seronegative sheep indicated that Hardjobovis seropositivity found in sheep may be able to predict shedding or renal carriage. © 2014 The Author(s).

Toward the Standardization of Biochar Analysis: The COST Action TD1107 Interlaboratory Comparison.

PubMed

Bachmann, Hans Jörg; Bucheli, Thomas D; Dieguez-Alonso, Alba; Fabbri, Daniele; Knicker, Heike; Schmidt, Hans-Peter; Ulbricht, Axel; Becker, Roland; Buscaroli, Alessandro; Buerge, Diane; Cross, Andrew; Dickinson, Dane; Enders, Akio; Esteves, Valdemar I; Evangelou, Michael W H; Fellet, Guido; Friedrich, Kevin; Gasco Guerrero, Gabriel; Glaser, Bruno; Hanke, Ulrich M; Hanley, Kelly; Hilber, Isabel; Kalderis, Dimitrios; Leifeld, Jens; Masek, Ondrej; Mumme, Jan; Carmona, Marina Paneque; Calvelo Pereira, Roberto; Rees, Frederic; Rombolà, Alessandro G; de la Rosa, José Maria; Sakrabani, Ruben; Sohi, Saran; Soja, Gerhard; Valagussa, Massimo; Verheijen, Frank; Zehetner, Franz

2016-01-20

Biochar produced by pyrolysis of organic residues is increasingly used for soil amendment and many other applications. However, analytical methods for its physical and chemical characterization are yet far from being specifically adapted, optimized, and standardized. Therefore, COST Action TD1107 conducted an interlaboratory comparison in which 22 laboratories from 12 countries analyzed three different types of biochar for 38 physical-chemical parameters (macro- and microelements, heavy metals, polycyclic aromatic hydrocarbons, pH, electrical conductivity, and specific surface area) with their preferential methods. The data were evaluated in detail using professional interlaboratory testing software. Whereas intralaboratory repeatability was generally good or at least acceptable, interlaboratory reproducibility was mostly not (20% < mean reproducibility standard deviation < 460%). This paper contributes to better comparability of biochar data published already and provides recommendations to improve and harmonize specific methods for biochar analysis in the future.

Forensic analysis of explosives using isotope ratio mass spectrometry (IRMS)--part 2: forensic inter-laboratory trial: bulk carbon and nitrogen stable isotopes in a range of chemical compounds (Australia and New Zealand).

PubMed

Benson, Sarah J; Lennard, Christopher J; Maynard, Philip; Hill, David M; Andrew, Anita S; Neal, Ken; Stuart-Williams, Hilary; Hope, Janet; Walker, G Stewart; Roux, Claude

2010-01-01

Comparability of data over time and between laboratories is a key issue for consideration in the development of global databases, and more broadly for quality assurance in general. One mechanism that can be utilized for evaluating traceability is an inter-laboratory trial. This paper addresses an inter-laboratory trial conducted across a number of Australian and New Zealand isotope ratio mass spectrometry (IRMS) laboratories. The main objective of this trial was to determine whether IRMS laboratories in these countries would record comparable values for the distributed samples. Four carbon containing and four nitrogen containing compounds were distributed to seven laboratories in Australia and one in New Zealand. The laboratories were requested to analyze the samples using their standard procedures. The data from each laboratory was evaluated collectively using International Standard ISO 13528 (Statistical methods for use in proficiency testing by inter-laboratory comparisons). "Warning signals" were raised against one participant in this trial. "Action signals" requiring corrective action were raised against four participants. These participants reviewed the data and possible sources for the discrepancies. This inter-laboratory trial was successful in providing an initial snapshot of the potential for traceability between the participating laboratories. The statistical methods described in this article could be used as a model for others needing to evaluate stable isotope results derived from multiple laboratories, e.g., inter-laboratory trials/proficiency testing. Ongoing trials will be conducted to improve traceability across the Australian and New Zealand IRMS community.

United Nations Environment Programme Capacity Building Pilot Project--training and interlaboratory study on persistent organic pollutant analysis under the Stockholm Convention.

PubMed

de Boer, J; Leslie, H; van Leeuwen, S P J; Wegener, J-W; van Bavel, B; Lindström, G; Lahoutifard, N; Fiedler, H

2008-06-09

Within the framework of a United Nations Environment Programme (UNEP) Capacity Building Project for training of laboratory staff in developing countries on persistent organic pollutant (POP) analysis, an interlaboratory study was organised following an initial evaluation of the performance of laboratories (reality check) and a series of training sessions. The target compounds were polychlorinated biphenyls (PCB) and organochlorine pesticides (OCP). Seven laboratories from five countries (Ecuador, Uruguay, Kenya, Moldova, and Fiji) participated. Most of the laboratories had no experience in determining PCBs. Although chromatograms improved considerably after the training and installation of new gas chromatographic (GC) columns at participating laboratories, the level of performance in the interlaboratory study was essentially on par with the moderate performance level achieved by European POP laboratories in the 1980s. Only some individual results were within +/-20% of the target values. The relative standard deviations (R.S.D.s) in POP concentrations determined by laboratories in a sediment sample were >200% in a number of cases. The results for a certified herring sample were better with at least some R.S.D. values below 50% and most below 100%. Clean up was as one of the main sources of error. After inspection it was ascertained that training of laboratory staff and investments in simple consumables such as glassware and GC columns would help to improve the quality of the analysis more than major investments in expensive instrumentation. Creating an effective network of POP laboratories at different continents together with a series of interlaboratory studies and workshops is suggested to improve the measurements of POPs in these countries.

[Interlaboratory Study on Evaporation Residue Test for Food Contact Products (Report 2)].

PubMed

Ohno, Hiroyuki; Mutsuga, Motoh; Abe, Tomoyuki; Abe, Yutaka; Amano, Homare; Ishihara, Kinuyo; Ohsaka, Ikue; Ohno, Haruka; Ohno, Yuichiro; Ozaki, Asako; Kakihara, Yoshiteru; Kobayashi, Hisashi; Sakuragi, Hiroshi; Shibata, Hiroshi; Shirono, Katsuhiro; Sekido, Haruko; Takasaka, Noriko; Takenaka, Yu; Tajima, Yoshiyasu; Tanaka, Aoi; Tanaka, Hideyuki; Nakanishi, Toru; Nomura, Chie; Haneishi, Nahoko; Hayakawa, Masato; Miura, Toshihiko; Yamaguchi, Miku; Yamada, Kyohei; Watanabe, Kazunari; Sato, Kyoko

2018-01-01

An interlaboratory study was performed to evaluate the equivalence between an official method and a modified method of evaporation residue test using heptane as a food-simulating solvent for oily or fatty foods, based on the Japanese Food Sanitation Law for food contact products. Twenty-three laboratories participated, and tested the evaporation residues of nine test solutions as blind duplicates. In the official method, heating for evaporation was done with a water bath. In the modified method, a hot plate was used for evaporation, and/or a vacuum concentration procedure was skipped. In most laboratories, the test solutions were heated until just prior to dryness, and then allowed to dry under residual heat. Statistical analysis revealed that there was no significant difference between the two methods. Accordingly, the modified method provides performance equal to the official method, and is available as an alternative method. Furthermore, an interlaboratory study was performed to evaluate and compare two leaching solutions (95% ethanol and isooctane) used as food-simulating solvents for oily or fatty foods in the EU. The results demonstrated that there was no significant difference between heptane and these two leaching solutions.

Results of an interlaboratory method performance study for the size determination and quantification of silver nanoparticles in chicken meat by single-particle inductively coupled plasma mass spectrometry (sp-ICP-MS).

PubMed

Weigel, Stefan; Peters, Ruud; Loeschner, Katrin; Grombe, Ringo; Linsinger, Thomas P J

2017-08-01

Single-particle inductively coupled plasma mass spectrometry (sp-ICP-MS) promises fast and selective determination of nanoparticle size and number concentrations. While several studies on practical applications have been published, data on formal, especially interlaboratory validation of sp-ICP-MS, is sparse. An international interlaboratory study was organized to determine repeatability and reproducibility of the determination of the median particle size and particle number concentration of Ag nanoparticles (AgNPs) in chicken meat. Ten laboratories from the European Union, the USA, and Canada determined particle size and particle number concentration of two chicken meat homogenates spiked with polyvinylpyrrolidone (PVP)-stabilized AgNPs. For the determination of the median particle diameter, repeatability standard deviations of 2 and 5% were determined, and reproducibility standard deviations were 15 and 25%, respectively. The equivalent median diameter itself was approximately 60% larger than the diameter of the particles in the spiking solution. Determination of the particle number concentration was significantly less precise, with repeatability standard deviations of 7 and 18% and reproducibility standard deviations of 70 and 90%.

Experimental design of an interlaboratory study for trace metal analysis of liquid fluids. [for aerospace vehicles

NASA Technical Reports Server (NTRS)

Greenbauer-Seng, L. A.

1983-01-01

The accurate determination of trace metals and fuels is an important requirement in much of the research into and development of alternative fuels for aerospace applications. Recognizing the detrimental effects of certain metals on fuel performance and fuel systems at the part per million and in some cases part per billion levels requires improved accuracy in determining these low concentration elements. Accurate analyses are also required to ensure interchangeability of analysis results between vendor, researcher, and end use for purposes of quality control. Previous interlaboratory studies have demonstrated the inability of different laboratories to agree on the results of metal analysis, particularly at low concentration levels, yet typically good precisions are reported within a laboratory. An interlaboratory study was designed to gain statistical information about the sources of variation in the reported concentrations. Five participant laboratories were used on a fee basis and were not informed of the purpose of the analyses. The effects of laboratory, analytical technique, concentration level, and ashing additive were studied in four fuel types for 20 elements of interest. The prescribed sample preparation schemes (variations of dry ashing) were used by all of the laboratories. The analytical data were statistically evaluated using a computer program for the analysis of variance technique.

Interlaboratory ring trial to evaluate CFT proficiency of European laboratories for diagnosis of glanders in equids.

PubMed

Laroucau, K; Colaneri, C; Jaÿ, M; Corde, Y; Drapeau, A; Durand, B; Zientara, S; Beck, C

2016-06-18

To evaluate the routine complement fixation test (CFT) used to detect Burkholderia mallei antibodies in equine sera, an interlaboratory proficiency test was held with 24 European laboratories, including 22 National Reference Laboratories for glanders. The panels sent to participants were composed of sera with or without B mallei antibodies. This study confirmed the reliability of CFT and highlighted its intralaboratory reproducibility. However, the sensitivity of glanders serodiagnosis and laboratory proficiency may be improved by standardising critical reagents, including antigens, and by developing a standard B mallei serum. British Veterinary Association.

Interlaboratory evaluation of a standardized inductively coupled plasma mass spectrometry method for the determination of trace beryllium in air filter samples.

PubMed

Ashley, Kevin; Brisson, Michael J; Howe, Alan M; Bartley, David L

2009-12-01

A collaborative interlaboratory evaluation of a newly standardized inductively coupled plasma mass spectrometry (ICP-MS) method for determining trace beryllium in workplace air samples was carried out toward fulfillment of method validation requirements for ASTM International voluntary consensus standard test methods. The interlaboratory study (ILS) was performed in accordance with an applicable ASTM International standard practice, ASTM E691, which describes statistical procedures for investigating interlaboratory precision. Uncertainty was also estimated in accordance with ASTM D7440, which applies the International Organization for Standardization Guide to the Expression of Uncertainty in Measurement to air quality measurements. Performance evaluation materials (PEMs) used consisted of 37 mm diameter mixed cellulose ester filters that were spiked with beryllium at levels of 0.025 (low loading), 0.5 (medium loading), and 10 (high loading) microg Be/filter; these spiked filters were prepared by a contract laboratory. Participating laboratories were recruited from a pool of over 50 invitees; ultimately, 20 laboratories from Europe, North America, and Asia submitted ILS results. Triplicates of each PEM (blanks plus the three different loading levels) were conveyed to each volunteer laboratory, along with a copy of the draft standard test method that each participant was asked to follow; spiking levels were unknown to the participants. The laboratories were requested to prepare the PEMs by one of three sample preparation procedures (hotplate or microwave digestion or hotblock extraction) that were described in the draft standard. Participants were then asked to analyze aliquots of the prepared samples by ICP-MS and to report their data in units of mu g Be/filter sample. Interlaboratory precision estimates from participating laboratories, computed in accordance with ASTM E691, were 0.165, 0.108, and 0.151 (relative standard deviation) for the PEMs spiked at 0.025, 0.5, and 10 microg Be/filter, respectively. Overall recoveries were 93.2%, 102%, and 80.6% for the low, medium, and high beryllium loadings, respectively. Expanded uncertainty estimates for interlaboratory analysis of low, medium, and high beryllium loadings, calculated in accordance with ASTM D7440, were 18.8%, 19.8%, and 24.4%, respectively. These figures of merit support promulgation of the analytical procedure as an ASTM International standard test method, ASTM D7439.

INTERLABORATORY COMPARISON OF A REDUCED VOLUME MARINE SEDIMENT TOXICITY TEST METHOD USING AMPHIPOD AMPELISCA ABDITA

EPA Science Inventory

The U.S. Environmental Protection Agency has standardized methods for performing acute marine amphipod sediment toxicity tests. A test design reducing sediment volume from 200 to 50 ml and overlying water from 600 to 150 ml was recently proposed. An interlaboratory comparison wa...

Extension of the validation of AOAC Official Method 2005.06 for dc-GTX2,3: interlaboratory study.

PubMed

Ben-Gigirey, Begoña; Rodríguez-Velasco, María L; Gago-Martínez, Ana

2012-01-01

AOAC Official Method(SM) 2005.06 for the determination of saxitoxin (STX)-group toxins in shellfish by LC with fluorescence detection with precolumn oxidation was previously validated and adopted First Action following a collaborative study. However, the method was not validated for all key STX-group toxins, and procedures to quantify some of them were not provided. With more STX-group toxin standards commercially available and modifications to procedures, it was possible to overcome some of these difficulties. The European Union Reference Laboratory for Marine Biotoxins conducted an interlaboratory exercise to extend AOAC Official Method 2005.06 validation for dc-GTX2,3 and to compile precision data for several STX-group toxins. This paper reports the study design and the results obtained. The performance characteristics for dc-GTX2,3 (intralaboratory and interlaboratory precision, recovery, and theoretical quantification limit) were evaluated. The mean recoveries obtained for dc-GTX2,3 were, in general, low (53.1-58.6%). The RSD for reproducibility (RSD(r)%) for dc-GTX2,3 in all samples ranged from 28.2 to 45.7%, and HorRat values ranged from 1.5 to 2.8. The article also describes a hydrolysis protocol to convert GTX6 to NEO, which has been proven to be useful for the quantification of GTX6 while the GTX6 standard is not available. The performance of the participant laboratories in the application of this method was compared with that obtained from the original collaborative study of the method. Intralaboratory and interlaboratory precision data for several STX-group toxins, including dc-NEO and GTX6, are reported here. This study can be useful for those laboratories determining STX-group toxins to fully implement AOAC Official Method 2005.06 for official paralytic shellfish poisoning control. However the overall quantitative performance obtained with the method was poor for certain toxins.

Centrally Determined Standardization of Flow Cytometry Methods Reduces Interlaboratory Variation in a Prospective Multicenter Study.

PubMed

Westera, Liset; van Viegen, Tanja; Jeyarajah, Jenny; Azad, Azar; Bilsborough, Janine; van den Brink, Gijs R; Cremer, Jonathan; Danese, Silvio; D'Haens, Geert; Eckmann, Lars; Faubion, William; Filice, Melissa; Korf, Hannelie; McGovern, Dermot; Panes, Julian; Salas, Azucena; Sandborn, William J; Silverberg, Mark S; Smith, Michelle I; Vermeire, Severine; Vetrano, Stefania; Shackelton, Lisa M; Stitt, Larry; Jairath, Vipul; Levesque, Barrett G; Spencer, David M; Feagan, Brian G; Vande Casteele, Niels

2017-11-02

Flow cytometry (FC) aids in characterization of cellular and molecular factors involved in pathologic immune responses. Although FC has potential to facilitate early drug development in inflammatory bowel disease, interlaboratory variability limits its use in multicenter trials. Standardization of methods may address this limitation. We compared variability in FC-aided quantitation of T-cell responses across international laboratories using three analytical strategies. Peripheral blood mononuclear cells (PBMCs) were isolated from three healthy donors, stimulated with phorbol 12-myristate 13-acetate and ionomycin at a central laboratory, fixed, frozen, and shipped to seven international laboratories. Permeabilization and staining was performed in triplicate at each laboratory using a common protocol and centrally provided reagents. Gating was performed using local gating with a local strategy (LGLS), local gating with a central strategy (LGCS), and central gating (CG). Median cell percentages were calculated across triplicates and donors, and reported for each condition and strategy. The coefficient of variation (CV) was calculated across laboratories. Between-strategy comparisons were made using a two-way analysis of variance adjusting for donor. Mean interlaboratory CV ranged from 1.8 to 102.1% depending on cell population and gating strategy (LGLS, 4.4-102.1%; LGCS, 10.9-65.6%; CG, 1.8-20.9%). Mean interlaboratory CV differed significantly across strategies and was consistently lower with CG. Central gating was the only strategy with mean CVs consistently lower than 25%, which is a proposed standard for pharmacodynamic and exploratory biomarker assays.

Intra- and interlaboratory variability in acute toxicity tests with glochidia and juveniles of freshwater mussels (Unionidae)

USGS Publications Warehouse

Wang, N.; Augspurger, T.; Barnhart, M.C.; Bidwell, Joseph R.; Cope, W.G.; Dwyer, F.J.; Geis, S.; Greer, I.E.; Ingersoll, C.G.; Kane, C.M.; May, T.W.; Neves, R.J.; Newton, T.J.; Roberts, A.D.; Whites, D.W.

2007-01-01

The present study evaluated the performance and variability in acute toxicity tests with glochidia and newly transformed juvenile mussels using the standard methods outlined in American Society for Testing and Materials (ASTM). Multiple 48-h toxicity tests with glochidia and 96-h tests with juvenile mussels were conducted within a single laboratory and among five laboratories. All tests met the test acceptability requirements (e.g., ???90% control survival). Intralaboratory tests were conducted over two consecutive mussel-spawning seasons with mucket (Actinonaias ligamentina) or fatmucket (Lampsilis siliquoidea) using copper, ammonia, or chlorine as a toxicant. For the glochidia of both species, the variability of intralaboratory median effective concentrations (EC50s) for the three toxicants, expressed as the coefficient of variation (CV), ranged from 14 to 27% in 24-h exposures and from 13 to 36% in 48-h exposures. The intralaboratory CV of copper EC50s for juvenile fatmucket was 24% in 48-h exposures and 13% in 96-h exposures. Interlaboratory tests were conducted with fatmucket glochidia and juveniles by five laboratories using copper as a toxicant. The interlaboratory CV of copper EC50s for glochidia was 13% in 24-h exposures and 24% in 48-h exposures, and the interlaboratory CV for juveniles was 22% in 48-h exposures and 42% in 96-h exposures. The high completion success and the overall low variability in test results indicate that the test methods have acceptable precision and can be performed routinely. ?? 2007 SETAC.

Sample integrity evaluation and EPA method 325B interlaboratory comparison for select volatile organic compounds collected diffusively on Carbopack X sorbent tubes

NASA Astrophysics Data System (ADS)

Oliver, Karen D.; Cousett, Tamira A.; Whitaker, Donald A.; Smith, Luther A.; Mukerjee, Shaibal; Stallings, Casson; Thoma, Eben D.; Alston, Lillian; Colon, Maribel; Wu, Tai; Henkle, Stacy

2017-08-01

A sample integrity evaluation and an interlaboratory comparison were conducted in application of U.S. Environmental Protection Agency (EPA) Methods 325A and 325B for diffusively monitoring benzene and other selected volatile organic compounds (VOCs) using Carbopack X sorbent tubes. To evaluate sample integrity, VOC samples were refrigerated for up to 240 days and analyzed using thermal desorption/gas chromatography-mass spectrometry at the EPA Office of Research and Development laboratory in Research Triangle Park, NC, USA. For the interlaboratory comparison, three commercial analytical laboratories were asked to follow Method 325B when analyzing samples of VOCs that were collected in field and laboratory settings for EPA studies. Overall results indicate that the selected VOCs collected diffusively on sorbent tubes generally were stable for 6 months or longer when samples were refrigerated. This suggests the specified maximum 30-day storage time of VOCs collected diffusively on Carbopack X passive samplers and analyzed using Method 325B might be able to be relaxed. Interlaboratory comparison results were in agreement for the challenge samples collected diffusively in an exposure chamber in the laboratory, with most measurements within ±25% of the theoretical concentration. Statistically significant differences among laboratories for ambient challenge samples were small, less than 1 part per billion by volume (ppbv). Results from all laboratories exhibited good precision and generally agreed well with each other.

Interlaboratory Reproducibility and Proficiency Testing within the Human Papillomavirus Cervical Cancer Screening Program in Catalonia, Spain

PubMed Central

Ibáñez, R.; Félez-Sánchez, M.; Godínez, J. M.; Guardià, C.; Caballero, E.; Juve, R.; Combalia, N.; Bellosillo, B.; Cuevas, D.; Moreno-Crespi, J.; Pons, L.; Autonell, J.; Gutierrez, C.; Ordi, J.; de Sanjosé, S.

2014-01-01

In Catalonia, a screening protocol for cervical cancer, including human papillomavirus (HPV) DNA testing using the Digene Hybrid Capture 2 (HC2) assay, was implemented in 2006. In order to monitor interlaboratory reproducibility, a proficiency testing (PT) survey of the HPV samples was launched in 2008. The aim of this study was to explore the repeatability of the HC2 assay's performance. Participating laboratories provided 20 samples annually, 5 randomly chosen samples from each of the following relative light unit (RLU) intervals: <0.5, 0.5 to 0.99, 1 to 9.99, and ≥10. Kappa statistics were used to determine the agreement levels between the original and the PT readings. The nature and origin of the discrepant results were calculated by bootstrapping. A total of 946 specimens were retested. The kappa values were 0.91 for positive/negative categorical classification and 0.79 for the four RLU intervals studied. Sample retesting yielded systematically lower RLU values than the original test (P < 0.005), independently of the time elapsed between the two determinations (median, 53 days), possibly due to freeze-thaw cycles. The probability for a sample to show clinically discrepant results upon retesting was a function of the RLU value; samples with RLU values in the 0.5 to 5 interval showed 10.80% probability to yield discrepant results (95% confidence interval [CI], 7.86 to 14.33) compared to 0.85% probability for samples outside this interval (95% CI, 0.17 to 1.69). Globally, the HC2 assay shows high interlaboratory concordance. We have identified differential confidence thresholds and suggested the guidelines for interlaboratory PT in the future, as analytical quality assessment of HPV DNA detection remains a central component of the screening program for cervical cancer prevention. PMID:24574284

Inter-Laboratory Comparison for Calibration of Relative Humidity Devices Among Accredited Laboratories in Malaysia

NASA Astrophysics Data System (ADS)

Hussain, F.; Khairuddin, S.; Othman, H.

2017-01-01

An inter-laboratory comparison in relative humidity measurements among accredited laboratories has been coordinated by the National Metrology Institute of Malaysia. It was carried out to determine the performance of the participating laboratories. The objective of the comparison was to acknowledge the participating laboratories competencies and to verify the level of accuracies declared in their scope of accreditation, in accordance with the MS ISO/IEC 17025 accreditation. The measurement parameter involved was relative humidity for the range of 30-90 %rh at a nominal temperature of 50°C. Eight accredited laboratories participated in the inter-laboratory comparison. Two units of artifacts have been circulated among the participants as the transfer standards.

INTERLABORATORY COMPARISON OF MASS SPECTROMETRIC METHODS FOR LEAD ISOTOPES AND TRACE ELEMENTS IN NIST SRM 1400 BONE ASH

EPA Science Inventory

The results of an interlaboratory comparison are reported for he lead isotope composition and for trace element concentrations in NIST SRM 1400 Bone Ash obtained using quadrupole and magnetic-sector inductively coupled plasma mass spectrometry (ICP-MS) and (for the Pb isotopes on...

Precision of the upgraded cottonscan instrument for measuring the average fiber linear density (fineness) of cotton lint samples

USDA-ARS?s Scientific Manuscript database

An inter-laboratory trial was conducted to validate the operation of the CottonscanTM technology as useful technique for determining the average fiber linear density of cotton. A significant inter-laboratory trial was completed and confirmed that the technology is quite acceptable. For fibers fin...

Assessing variability in chemical acute toxicity of unionid mussels: Influence of intra- and inter-laboratory testing, life stage, and species

EPA Science Inventory

The authors developed a toxicity database for unionid mussels to examine the extent of intra- and interlaboratory variability in acute toxicity tests with mussel larvae (glochidia) and juveniles; the extent of differential sensitivity of the 2 life stages; and the variation in se...

Sample Integrity Evaluation and EPA Method 325b Interlaboratory Comparison for Select Volatile Organic Compounds Collected Diffusively on Carbopack X Sorbent Tubes

EPA Science Inventory

Sample integrity evaluations and inter-laboratory comparisons were conducted in application of U.S. Environmental Protection Agency (EPA) Methods 325A/B for monitoring benzene and additional selected volatile organic compounds (VOCs) usingpassive-diffusive Carbopack X tube sample...

A scoring metric for multivariate data for reproducibility analysis using chemometric methods

PubMed Central

Sheen, David A.; de Carvalho Rocha, Werickson Fortunato; Lippa, Katrice A.; Bearden, Daniel W.

2017-01-01

Process quality control and reproducibility in emerging measurement fields such as metabolomics is normally assured by interlaboratory comparison testing. As a part of this testing process, spectral features from a spectroscopic method such as nuclear magnetic resonance (NMR) spectroscopy are attributed to particular analytes within a mixture, and it is the metabolite concentrations that are returned for comparison between laboratories. However, data quality may also be assessed directly by using binned spectral data before the time-consuming identification and quantification. Use of the binned spectra has some advantages, including preserving information about trace constituents and enabling identification of process difficulties. In this paper, we demonstrate the use of binned NMR spectra to conduct a detailed interlaboratory comparison and composition analysis. Spectra of synthetic and biologically-obtained metabolite mixtures, taken from a previous interlaboratory study, are compared with cluster analysis using a variety of distance and entropy metrics. The individual measurements are then evaluated based on where they fall within their clusters, and a laboratory-level scoring metric is developed, which provides an assessment of each laboratory’s individual performance. PMID:28694553

Interlaboratory comparison for the determination of the soluble fraction of metals in welding fume samples.

PubMed

Berlinger, Balazs; Harper, Martin

2018-02-01

There is interest in the bioaccessible metal components of aerosols, but this has been minimally studied because standardized sampling and analytical methods have not yet been developed. An interlaboratory study (ILS) has been carried out to evaluate a method for determining the water-soluble component of realistic welding fume (WF) air samples. Replicate samples were generated in the laboratory and distributed to participating laboratories to be analyzed according to a standardized procedure. Within-laboratory precision of replicate sample analysis (repeatability) was very good. Reproducibility between laboratories was not as good, but within limits of acceptability for the analysis of typical aerosol samples. These results can be used to support the development of a standardized test method.

Interlaboratory quality control of total HIV-1 DNA load measurement for multicenter reservoir studies.

PubMed

Gantner, Pierre; Mélard, Adeline; Damond, Florence; Delaugerre, Constance; Dina, Julia; Gueudin, Marie; Maillard, Anne; Sauné, Karine; Rodallec, Audrey; Tuaillon, Edouard; Plantier, Jean-Christophe; Rouzioux, Christine; Avettand-Fenoel, Véronique

2017-11-01

Viral reservoirs represent an important barrier to HIV cure. Accurate markers of HIV reservoirs are needed to develop multicenter studies. The aim of this multicenter quality control (QC) was to evaluate the inter-laboratory reproducibility of total HIV-1-DNA quantification. Ten laboratories of the ANRS-AC11 working group participated by quantifying HIV-DNA with a real-time qPCR assay (Biocentric) in four samples (QCMD). Good reproducibility was found between laboratories (standard deviation ≤ 0.2 log 10 copies/10 6 PBMC) for the three positive QC that were correctly classified by each laboratory (QC1

Assessing variability in chemical acute toxicity of unionid mussels: Influence of intra- and inter-laboratory testing, life stage, and species - SETAC Abstract

EPA Science Inventory

We developed a toxicity database for unionid mussels to examine the extent of intra- and inter-laboratory variability in acute toxicity tests with mussel larvae (glochidia) and juveniles; the extent of differential sensitivity of the two life stages; and the variation in sensitiv...

Role and Evaluation of Interlaboratory Comparison Results in Laboratory Accreditation

NASA Astrophysics Data System (ADS)

Bode, P.

2008-08-01

Participation in interlaboratory comparisons provides laboratories an opportunity for independent assessment of their analytical performance, both in absolute way and in comparison with those by other techniques. However, such comparisons are hindered by differences in the way laboratories participate, e.g. at best measurement capability or under routine conditions. Neutron activation analysis laboratories, determining total mass fractions, often see themselves classified as `outliers' since the majority of other participants employ techniques with incomplete digestion methods. These considerations are discussed in relation to the way results from interlaboratory comparisons are evaluated by accreditation bodies following the requirements of Clause 5.9.1 of the ISO/IEC 17025:2005. The discussion and conclusions come largely forth from experiences in the author's own laboratory.

Centrally Determined Standardization of Flow Cytometry Methods Reduces Interlaboratory Variation in a Prospective Multicenter Study

PubMed Central

Westera, Liset; van Viegen, Tanja; Jeyarajah, Jenny; Azad, Azar; Bilsborough, Janine; van den Brink, Gijs R; Cremer, Jonathan; Danese, Silvio; D'Haens, Geert; Eckmann, Lars; Faubion, William; Filice, Melissa; Korf, Hannelie; McGovern, Dermot; Panes, Julian; Salas, Azucena; Sandborn, William J; Silverberg, Mark S; Smith, Michelle I; Vermeire, Severine; Vetrano, Stefania; Shackelton, Lisa M; Stitt, Larry; Jairath, Vipul; Levesque, Barrett G; Spencer, David M; Feagan, Brian G; Vande Casteele, Niels

2017-01-01

Objectives: Flow cytometry (FC) aids in characterization of cellular and molecular factors involved in pathologic immune responses. Although FC has potential to facilitate early drug development in inflammatory bowel disease, interlaboratory variability limits its use in multicenter trials. Standardization of methods may address this limitation. We compared variability in FC-aided quantitation of T-cell responses across international laboratories using three analytical strategies. Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from three healthy donors, stimulated with phorbol 12-myristate 13-acetate and ionomycin at a central laboratory, fixed, frozen, and shipped to seven international laboratories. Permeabilization and staining was performed in triplicate at each laboratory using a common protocol and centrally provided reagents. Gating was performed using local gating with a local strategy (LGLS), local gating with a central strategy (LGCS), and central gating (CG). Median cell percentages were calculated across triplicates and donors, and reported for each condition and strategy. The coefficient of variation (CV) was calculated across laboratories. Between-strategy comparisons were made using a two-way analysis of variance adjusting for donor. Results: Mean interlaboratory CV ranged from 1.8 to 102.1% depending on cell population and gating strategy (LGLS, 4.4–102.1% LGCS, 10.9–65.6% CG, 1.8–20.9%). Mean interlaboratory CV differed significantly across strategies and was consistently lower with CG. Conclusions: Central gating was the only strategy with mean CVs consistently lower than 25%, which is a proposed standard for pharmacodynamic and exploratory biomarker assays. PMID:29095427

An Interlaboratory Evaluation of Drift Tube Ion Mobility–Mass Spectrometry Collision Cross Section Measurements

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stow, Sarah M.; Causon, Tim J.; Zheng, Xueyun

Collision cross section (CCS) measurements resulting from ion mobility-mass spectrometry (IM-MS) experiments provide a promising orthogonal dimension of structural information in MS-based analytical separations. As with any molecular identifier, interlaboratory standardization must precede broad range integration into analytical workflows. In this study, we present a reference drift tube ion mobility mass spectrometer (DTIM-MS) where improvements on the measurement accuracy of experimental parameters influencing IM separations provide standardized drift tube, nitrogen CCS values (DTCCSN2) for over 120 unique ion species with the lowest measurement uncertainty to date. The reproducibility of these DTCCSN2 values are evaluated across three additional laboratories on amore » commercially available DTIM-MS instrument. The traditional stepped field CCS method performs with a relative standard deviation (RSD) of 0.29% for all ion species across the three additional laboratories. The calibrated single field CCS method, which is compatible with a wide range of chromatographic inlet systems, performs with an average, absolute bias of 0.54% to the standardized stepped field DTCCSN2 values on the reference system. The low RSD and biases observed in this interlaboratory study illustrate the potential of DTIM-MS for providing a molecular identifier for a broad range of discovery based analyses.« less

Advancing the Use of Passive Sampling in Risk Assessment and Management of Sediments Contaminated with Hydrophobic Organic Chemicals: Results of an International Ex Situ Passive Sampling Interlaboratory Comparison

EPA Science Inventory

This work presents the results of an international interlaboratory comparison on ex situ passive sampling in sediments. The main objectives were to map the state of the science in passively sampling sediments, identify sources of variability, provide recommendations and practica...

Advancing the Use of Passive Sampling in Risk Assessment and Management of Sediments Contaminated with Hydrophobic Organic Chemicals: Results of an International Ex Situ Passive Sampling Interlaboratory Comparison

EPA Science Inventory

This work presents the results of an international interlaboratory comparison on ex situ passive sampling in sediments. The main objectives were to map the state of the science in passively sampling sediments, identify sources of variability, provide recommendations and practical...

Empirical insights and considerations for the OBT inter-laboratory comparison of environmental samples.

PubMed

Kim, Sang-Bog; Roche, Jennifer

2013-08-01

Organically bound tritium (OBT) is an important tritium species that can be measured in most environmental samples, but has only recently been recognized as a species of tritium in these samples. Currently, OBT is not routinely measured by environmental monitoring laboratories around the world. There are no certified reference materials (CRMs) for environmental samples. Thus, quality assurance (QA), or verification of the accuracy of the OBT measurement, is not possible. Alternatively, quality control (QC), or verification of the precision of the OBT measurement, can be achieved. In the past, there have been differences in OBT analysis results between environmental laboratories. A possible reason for the discrepancies may be differences in analytical methods. Therefore, inter-laboratory OBT comparisons among the environmental laboratories are important and would provide a good opportunity for adopting a reference OBT analytical procedure. Due to the analytical issues, only limited information is available on OBT measurement. Previously conducted OBT inter-laboratory practices are reviewed and the findings are described. Based on our experiences, a few considerations were suggested for the international OBT inter-laboratory comparison exercise to be completed in the near future. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

Development and Interlaboratory Study of a Liquid Chromatography Tandem Mass Spectrometric Methodfor the Determination of Multiple Mycotoxins inCereals Using Stable Isotope Dilution.

PubMed

Ye, Jin; Wu, Yu; Guo, Qilei; Lu, Meiling; Wang, Songshan; Xin, Yuanyuan; Xie, Gang; Zhang, Yan; Mariappan, Meena; Wang, Songxue

2018-05-01

An efficient, rapid, accurate, and cost-effective method based on stable isotope dilution and LC tandem MS was developed for the determination of multimycotoxins in cereals. The samples were extracted using acetonitrile-water-acetic acid (70 + 29 + 1, v/v/v), followed by dilution and centrifugation without any further cleanup. The mycotoxins were separated on a C18 column. Interference due to matrix effects was efficiently compensated for with [13C]-labeled stable isotope internal standards. The method demonstrated excellent linear relations, with regression coefficients above 0.999. Spiked recoveries at three different concentrations ranged from 80.9 to 115.9%, and RSDs were below 14% for all mycotoxins. The trueness of the method was also verified by participating in two proficiency tests, and satisfactory z-scores (|z| < 1.1) were obtained. In addition, an international interlaboratory study was organized to evaluate the methods. Eight laboratories characterized recovery, repeatability, and reproducibility studies in wheat, maize, and barley. The interlaboratory results were analyzed according to ISO 5725-2. Cochran and Grubbs tests were used to remove outliers. The mean recoveries of all 16 mycotoxins ranged from 87 to 111%. Repeatability, reproducibility, and Horwitz ratio values were 3.5-16.2, 5.4-33.6, and 0.16-1.65%, respectively. The results demonstrate that the method is reliable to determine multimycotoxins in cereals.

Measuring the uncertainties of discharge measurements: interlaboratory experiments in hydrometry

NASA Astrophysics Data System (ADS)

Le Coz, Jérôme; Blanquart, Bertrand; Pobanz, Karine; Dramais, Guillaume; Pierrefeu, Gilles; Hauet, Alexandre; Despax, Aurélien

2015-04-01

Quantifying the uncertainty of streamflow data is key for hydrological sciences. The conventional uncertainty analysis based on error propagation techniques is restricted by the absence of traceable discharge standards and by the weight of difficult-to-predict errors related to the operator, procedure and measurement environment. Field interlaboratory experiments recently emerged as an efficient, standardized method to 'measure' the uncertainties of a given streamgauging technique in given measurement conditions. Both uncertainty approaches are compatible and should be developed jointly in the field of hydrometry. In the recent years, several interlaboratory experiments have been reported by different hydrological services. They involved different streamgauging techniques, including acoustic profilers (ADCP), current-meters and handheld radars (SVR). Uncertainty analysis was not always their primary goal: most often, testing the proficiency and homogeneity of instruments, makes and models, procedures and operators was the original motivation. When interlaboratory experiments are processed for uncertainty analysis, once outliers have been discarded all participants are assumed to be equally skilled and to apply the same streamgauging technique in equivalent conditions. A universal requirement is that all participants simultaneously measure the same discharge, which shall be kept constant within negligible variations. To our best knowledge, we were the first to apply the interlaboratory method for computing the uncertainties of streamgauging techniques, according to the authoritative international documents (ISO standards). Several specific issues arise due to the measurements conditions in outdoor canals and rivers. The main limitation is that the best available river discharge references are usually too uncertain to quantify the bias of the streamgauging technique, i.e. the systematic errors that are common to all participants in the experiment. A reference or a sensitivity analysis to the fixed parameters of the streamgauging technique remain very useful for estimating the uncertainty related to the (non quantified) bias correction. In the absence of a reference, the uncertainty estimate is referenced to the average of all discharge measurements in the interlaboratory experiment, ignoring the technique bias. Simple equations can be used to assess the uncertainty of the uncertainty results, as a function of the number of participants and of repeated measurements. The interlaboratory method was applied to several interlaboratory experiments on ADCPs and currentmeters mounted on wading rods, in streams of different sizes and aspects, with 10 to 30 instruments, typically. The uncertainty results were consistent with the usual expert judgment and highly depended on the measurement environment. Approximately, the expanded uncertainties (within the 95% probability interval) were ±5% to ±10% for ADCPs in good or poor conditions, and ±10% to ±15% for currentmeters in shallow creeks. Due to the specific limitations related to a slow measurement process and to small, natural streams, uncertainty results for currentmeters were more uncertain than for ADCPs, for which the site-specific errors were significantly evidenced. The proposed method can be applied to a wide range of interlaboratory experiments conducted in contrasted environments for different streamgauging techniques, in a standardized way. Ideally, an international open database would enhance the investigation of hydrological data uncertainties, according to the characteristics of the measurement conditions and procedures. Such a dataset could be used for implementing and validating uncertainty propagation methods in hydrometry.

Rapid Analysis of Carbohydrates in Bioprocess Samples: An Evaluation of the CarboPac SA10 for HPAE-PAD Analysis by Interlaboratory Comparison

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sevcik, R. S.; Hyman, D. A.; Basumallich, L.

2013-01-01

A technique for carbohydrate analysis for bioprocess samples has been developed, providing reduced analysis time compared to current practice in the biofuels R&D community. The Thermofisher CarboPac SA10 anion-exchange column enables isocratic separation of monosaccharides, sucrose and cellobiose in approximately 7 minutes. Additionally, use of a low-volume (0.2 mL) injection valve in combination with a high-volume detection cell minimizes the extent of sample dilution required to bring sugar concentrations into the linear range of the pulsed amperometric detector (PAD). Three laboratories, representing academia, industry, and government, participated in an interlaboratory study which analyzed twenty-one opportunistic samples representing biomass pretreatment, enzymaticmore » saccharification, and fermentation samples. The technique's robustness, linearity, and interlaboratory reproducibility were evaluated and showed excellent-to-acceptable characteristics. Additionally, quantitation by the CarboPac SA10/PAD was compared with the current practice method utilizing a HPX-87P/RID. While these two methods showed good agreement a statistical comparison found significant quantitation difference between them, highlighting the difference between selective and universal detection modes.« less

Interlaboratory comparison of three microbial source tracking quantitative polymerase chain reaction (qPCR) assays from fecal-source and environmental samples

USGS Publications Warehouse

Stelzer, Erin A.; Strickler, Kriston M.; Schill, William B.

2012-01-01

During summer and early fall 2010, 15 river samples and 6 fecal-source samples were collected in West Virginia. These samples were analyzed by three laboratories for three microbial source tracking (MST) markers: AllBac, a general fecal indicator; BacHum, a human-associated fecal indicator; and BoBac, a ruminant-associated fecal indicator. MST markers were analyzed by means of the quantitative polymerase chain reaction (qPCR) method. The aim was to assess interlaboratory precision when the three laboratories used the same MST marker and shared deoxyribonucleic acid (DNA) extracts of the samples, but different equipment, reagents, and analyst experience levels. The term assay refers to both the markers and the procedure differences listed above. Interlaboratory precision was best for all three MST assays when using the geometric mean absolute relative percent difference (ARPD) and Friedman's statistical test as a measure of interlaboratory precision. Adjustment factors (one for each MST assay) were calculated using results from fecal-source samples analyzed by all three laboratories and applied retrospectively to sample concentrations to account for differences in qPCR results among labs using different standards and procedures. Following the application of adjustment factors to qPCR results, ARPDs were lower; however, statistically significant differences between labs were still observed for the BacHum and BoBac assays. This was a small study and two of the MST assays had 52 percent of samples with concentrations at or below the limit of accurate quantification; hence, more testing could be done to determine if the adjustment factors would work better if the majority of sample concentrations were above the quantification limit.

Particle size distributions by transmission electron microscopy: an interlaboratory comparison case study

PubMed Central

Rice, Stephen B; Chan, Christopher; Brown, Scott C; Eschbach, Peter; Han, Li; Ensor, David S; Stefaniak, Aleksandr B; Bonevich, John; Vladár, András E; Hight Walker, Angela R; Zheng, Jiwen; Starnes, Catherine; Stromberg, Arnold; Ye, Jia; Grulke, Eric A

2015-01-01

This paper reports an interlaboratory comparison that evaluated a protocol for measuring and analysing the particle size distribution of discrete, metallic, spheroidal nanoparticles using transmission electron microscopy (TEM). The study was focused on automated image capture and automated particle analysis. NIST RM8012 gold nanoparticles (30 nm nominal diameter) were measured for area-equivalent diameter distributions by eight laboratories. Statistical analysis was used to (1) assess the data quality without using size distribution reference models, (2) determine reference model parameters for different size distribution reference models and non-linear regression fitting methods and (3) assess the measurement uncertainty of a size distribution parameter by using its coefficient of variation. The interlaboratory area-equivalent diameter mean, 27.6 nm ± 2.4 nm (computed based on a normal distribution), was quite similar to the area-equivalent diameter, 27.6 nm, assigned to NIST RM8012. The lognormal reference model was the preferred choice for these particle size distributions as, for all laboratories, its parameters had lower relative standard errors (RSEs) than the other size distribution reference models tested (normal, Weibull and Rosin–Rammler–Bennett). The RSEs for the fitted standard deviations were two orders of magnitude higher than those for the fitted means, suggesting that most of the parameter estimate errors were associated with estimating the breadth of the distributions. The coefficients of variation for the interlaboratory statistics also confirmed the lognormal reference model as the preferred choice. From quasi-linear plots, the typical range for good fits between the model and cumulative number-based distributions was 1.9 fitted standard deviations less than the mean to 2.3 fitted standard deviations above the mean. Automated image capture, automated particle analysis and statistical evaluation of the data and fitting coefficients provide a framework for assessing nanoparticle size distributions using TEM for image acquisition. PMID:26361398

Consensus building for interlaboratory studies, key comparisons, and meta-analysis

NASA Astrophysics Data System (ADS)

Koepke, Amanda; Lafarge, Thomas; Possolo, Antonio; Toman, Blaza

2017-06-01

Interlaboratory studies in measurement science, including key comparisons, and meta-analyses in several fields, including medicine, serve to intercompare measurement results obtained independently, and typically produce a consensus value for the common measurand that blends the values measured by the participants. Since interlaboratory studies and meta-analyses reveal and quantify differences between measured values, regardless of the underlying causes for such differences, they also provide so-called ‘top-down’ evaluations of measurement uncertainty. Measured values are often substantially over-dispersed by comparison with their individual, stated uncertainties, thus suggesting the existence of yet unrecognized sources of uncertainty (dark uncertainty). We contrast two different approaches to take dark uncertainty into account both in the computation of consensus values and in the evaluation of the associated uncertainty, which have traditionally been preferred by different scientific communities. One inflates the stated uncertainties by a multiplicative factor. The other adds laboratory-specific ‘effects’ to the value of the measurand. After distinguishing what we call recipe-based and model-based approaches to data reductions in interlaboratory studies, we state six guiding principles that should inform such reductions. These principles favor model-based approaches that expose and facilitate the critical assessment of validating assumptions, and give preeminence to substantive criteria to determine which measurement results to include, and which to exclude, as opposed to purely statistical considerations, and also how to weigh them. Following an overview of maximum likelihood methods, three general purpose procedures for data reduction are described in detail, including explanations of how the consensus value and degrees of equivalence are computed, and the associated uncertainty evaluated: the DerSimonian-Laird procedure; a hierarchical Bayesian procedure; and the Linear Pool. These three procedures have been implemented and made widely accessible in a Web-based application (NIST Consensus Builder). We illustrate principles, statistical models, and data reduction procedures in four examples: (i) the measurement of the Newtonian constant of gravitation; (ii) the measurement of the half-lives of radioactive isotopes of caesium and strontium; (iii) the comparison of two alternative treatments for carotid artery stenosis; and (iv) a key comparison where the measurand was the calibration factor of a radio-frequency power sensor.

Determination of the acid value of instant noodles: interlaboratory study.

PubMed

Hakoda, Akiko; Sakaida, Kenichi; Suzuki, Tadanao; Yasui, Akemi

2006-01-01

An interlaboratory study was performed to evaluate the method for determining the acid value of instant noodles, based on the Japanese Agricultural Standard (JAS), with extraction of lipid using petroleum ether at a volume of 100 mL to the test portion of 25 g. Thirteen laboratories participated and analyzed 5 test samples as blind duplicates. Statistical treatment revealed that the repeatability (RSDr) of acid value was <6.5%, and the reproducibility (RSDR) of acid value was <9.6%. The HorRat values (RSDR/predicted RSDR) were 1.2-1.8, where the RSDR and the predicted RSDR were obtained in terms of free fatty acids in the noodles per unit weight, using the equation [acid value = percent free fatty acids (as oleic) x 1.99] and the extracted lipid contents. This method was shown to have acceptable precision by the present study.

Bayesian Local Contamination Models for Multivariate Outliers

PubMed Central

Page, Garritt L.; Dunson, David B.

2013-01-01

In studies where data are generated from multiple locations or sources it is common for there to exist observations that are quite unlike the majority. Motivated by the application of establishing a reference value in an inter-laboratory setting when outlying labs are present, we propose a local contamination model that is able to accommodate unusual multivariate realizations in a flexible way. The proposed method models the process level of a hierarchical model using a mixture with a parametric component and a possibly nonparametric contamination. Much of the flexibility in the methodology is achieved by allowing varying random subsets of the elements in the lab-specific mean vectors to be allocated to the contamination component. Computational methods are developed and the methodology is compared to three other possible approaches using a simulation study. We apply the proposed method to a NIST/NOAA sponsored inter-laboratory study which motivated the methodological development. PMID:24363465

Variances and uncertainties of the sample laboratory-to-laboratory variance (S(L)2) and standard deviation (S(L)) associated with an interlaboratory study.

PubMed

McClure, Foster D; Lee, Jung K

2012-01-01

The validation process for an analytical method usually employs an interlaboratory study conducted as a balanced completely randomized model involving a specified number of randomly chosen laboratories, each analyzing a specified number of randomly allocated replicates. For such studies, formulas to obtain approximate unbiased estimates of the variance and uncertainty of the sample laboratory-to-laboratory (lab-to-lab) STD (S(L)) have been developed primarily to account for the uncertainty of S(L) when there is a need to develop an uncertainty budget that includes the uncertainty of S(L). For the sake of completeness on this topic, formulas to estimate the variance and uncertainty of the sample lab-to-lab variance (S(L)2) were also developed. In some cases, it was necessary to derive the formulas based on an approximate distribution for S(L)2.

Interlaboratory comparison of fluorocarbons-11, -12, methylchloroform and nitrous oxide measurements

NASA Technical Reports Server (NTRS)

Rasmussen, R. A.; Khalil, M. A. K.

1981-01-01

Measurements conducted by 19 participating laboratories were considered in the reported interlaboratory comparison study. The results show that there is considerable disagreement among laboratories regarding the absolute concentrations of all four trace gases (CCl3F, CCl2F2, H3CCl3, N2O). The magnitude of this disagreement is discussed. Laboratories in Group II showed considerable disagreement among themselves. Their results were scattered within large intervals of concentration. Laboratories in Group I (using common standards) were in excellent (+ or - 5%) agreement among themselves. A systematic disagreement was noted between Groups I and II laboratories. Generally, the mean values of concentrations determined from the measurements of Group II laboratories were lower than the mean values reported by Group I laboratories.

Evaluation of Brazilian Sugarcane Bagasse Characterization: An Interlaboratory Comparison Study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sluiter, Justin B.; Chum, Helena; Gomes, Absai C.

2016-05-01

This paper describes a study of the variability of measured composition for a single bulk sugarcane bagasse conducted across eight laboratories using similar analytical methods, with the purpose of determining the expected variation for compositional analysis performed by different laboratories. The results show good agreement of measured composition within a single laboratory, but greater variability when results are compared among laboratories. These interlaboratory variabilities do not seem to be associated with a specific method or technique or any single piece of instrumentation. The summary censored statistics provide mean values and pooled standard deviations as follows: total extractives 6.7% (0.6%), wholemore » ash 1.5% (0.2%), glucan 42.3% (1.2%), xylan 22.3% (0.5%), total lignin 21.3% (0.4%), and total mass closure 99.4% (2.9%).« less

[Interlaboratory Study on Evaporation Residue Test for Food Contact Products (Report 1)].

PubMed

Ohno, Hiroyuki; Mutsuga, Motoh; Abe, Tomoyuki; Abe, Yutaka; Amano, Homare; Ishihara, Kinuyo; Ohsaka, Ikue; Ohno, Haruka; Ohno, Yuichiro; Ozaki, Asako; Kakihara, Yoshiteru; Kobayashi, Hisashi; Sakuragi, Hiroshi; Shibata, Hiroshi; Shirono, Katsuhiro; Sekido, Haruko; Takasaka, Noriko; Takenaka, Yu; Tajima, Yoshiyasu; Tanaka, Aoi; Tanaka, Hideyuki; Tonooka, Hiroyuki; Nakanishi, Toru; Nomura, Chie; Haneishi, Nahoko; Hayakawa, Masato; Miura, Toshihiko; Yamaguchi, Miku; Watanabe, Kazunari; Sato, Kyoko

2018-01-01

An interlaboratory study was performed to evaluate the equivalence between an official method and a modified method of evaporation residue test using three food-simulating solvents (water, 4% acetic acid and 20% ethanol), based on the Japanese Food Sanitation Law for food contact products. Twenty-three laboratories participated, and tested the evaporation residues of nine test solutions as blind duplicates. For evaporation, a water bath was used in the official method, and a hot plate in the modified method. In most laboratories, the test solutions were heated until just prior to evaporation to dryness, and then allowed to dry under residual heat. Statistical analysis revealed that there was no significant difference between the two methods, regardless of the heating equipment used. Accordingly, the modified method provides performance equal to the official method, and is available as an alternative method.

First Worldwide Proficiency Study on Variable-Number Tandem-Repeat Typing of Mycobacterium tuberculosis Complex Strains

PubMed Central

de Beer, Jessica L.; Kremer, Kristin; Ködmön, Csaba; Supply, Philip

2012-01-01

Although variable-number tandem-repeat (VNTR) typing has gained recognition as the new standard for the DNA fingerprinting of Mycobacterium tuberculosis complex (MTBC) isolates, external quality control programs have not yet been developed. Therefore, we organized the first multicenter proficiency study on 24-locus VNTR typing. Sets of 30 DNAs of MTBC strains, including 10 duplicate DNA samples, were distributed among 37 participating laboratories in 30 different countries worldwide. Twenty-four laboratories used an in-house-adapted method with fragment sizing by gel electrophoresis or an automated DNA analyzer, nine laboratories used a commercially available kit, and four laboratories used other methods. The intra- and interlaboratory reproducibilities of VNTR typing varied from 0% to 100%, with averages of 72% and 60%, respectively. Twenty of the 37 laboratories failed to amplify particular VNTR loci; if these missing results were ignored, the number of laboratories with 100% interlaboratory reproducibility increased from 1 to 5. The average interlaboratory reproducibility of VNTR typing using a commercial kit was better (88%) than that of in-house-adapted methods using a DNA analyzer (70%) or gel electrophoresis (50%). Eleven laboratories using in-house-adapted manual typing or automated typing scored inter- and intralaboratory reproducibilities of 80% or higher, which suggests that these approaches can be used in a reliable way. In conclusion, this first multicenter study has documented the worldwide quality of VNTR typing of MTBC strains and highlights the importance of international quality control to improve genotyping in the future. PMID:22170917

Bias from two analytical laboratories involved in a long-term air monitoring program measuring organic pollutants in the Arctic: a quality assurance/quality control assessment.

PubMed

Su, Yushan; Hung, Hayley; Stern, Gary; Sverko, Ed; Lao, Randy; Barresi, Enzo; Rosenberg, Bruno; Fellin, Phil; Li, Henrik; Xiao, Hang

2011-11-01

Initiated in 1992, air monitoring of organic pollutants in the Canadian Arctic provided spatial and temporal trends in support of Canada's participation in the Stockholm Convention of Persistent Organic Pollutants. The specific analytical laboratory charged with this task was changed in 2002 while field sampling protocols remained unchanged. Three rounds of intensive comparison studies were conducted in 2004, 2005, and 2008 to assess data comparability between the two laboratories. Analysis was compared for organochlorine pesticides (OCPs), polychlorinated biphenyls (PCBs) and polycyclic aromatic hydrocarbons (PAHs) in standards, blind samples of mixed standards and extracts of real air samples. Good measurement accuracy was achieved for both laboratories when standards were analyzed. Variation of measurement accuracy over time was found for some OCPs and PCBs in standards on a random and non-systematic manner. Relatively low accuracy in analyzing blind samples was likely related to the process of sample purification. Inter-laboratory measurement differences for standards (<30%) and samples (<70%) were generally less than or comparable to those reported in a previous inter-laboratory study with 21 participating laboratories. Regression analysis showed inconsistent data comparability between the two laboratories during the initial stages of the study. These inter-laboratory differences can complicate abilities to discern long-term trends of pollutants in a given sampling site. It is advisable to maintain long-term measurements with minimal changes in sample analysis.

Validation of Dissolution Testing with Biorelevant Media: An OrBiTo Study.

PubMed

Mann, James; Dressman, Jennifer; Rosenblatt, Karin; Ashworth, Lee; Muenster, Uwe; Frank, Kerstin; Hutchins, Paul; Williams, James; Klumpp, Lukas; Wielockx, Kristina; Berben, Philippe; Augustijns, Patrick; Holm, Rene; Hofmann, Michael; Patel, Sanjaykumar; Beato, Stefania; Ojala, Krista; Tomaszewska, Irena; Bruel, Jean-Luc; Butler, James

2017-12-04

Dissolution testing with biorelevant media has become widespread in the pharmaceutical industry as a means of better understanding how drugs and formulations behave in the gastrointestinal tract. Until now, however, there have been few attempts to gauge the reproducibility of results obtained with these methods. The aim of this study was to determine the interlaboratory reproducibility of biorelevant dissolution testing, using the paddle apparatus (USP 2). Thirteen industrial and three academic laboratories participated in this study. All laboratories were provided with standard protocols for running the tests: dissolution in FaSSGF to simulate release in the stomach, dissolution in a single intestinal medium, FaSSIF, to simulate release in the small intestine, and a "transfer" (two-stage) protocol to simulate the concentration profile when conditions are changed from the gastric to the intestinal environment. The test products chosen were commercially available ibuprofen tablets and zafirlukast tablets. The biorelevant dissolution tests showed a high degree of reproducibility among the participating laboratories, even though several different batches of the commercially available medium preparation powder were used. Likewise, results were almost identicalbetween the commercial biorelevant media and those produced in-house. Comparing results to previous ring studies, including those performed with USP calibrator tablets or commercially available pharmaceutical products in a single medium, the results for the biorelevant studies were highly reproducible on an interlaboratory basis. Interlaboratory reproducibility with the two-stage test was also acceptable, although the variability was somewhat greater than with the single medium tests. Biorelevant dissolution testing is highly reproducible among laboratories and can be relied upon for cross-laboratory comparisons.

A SIMPLE METHOD FOR EVALUATING DATA FROM AN INTERLABORATORY STUDY

EPA Science Inventory

Large-scale laboratory-and method-performance studies involving more than about 30 laboratories may be evaluated by calculating the HORRAT ratio for each test sample (HORRAT=[experimentally found among-laboratories relative standard deviation] divided by [relative standard deviat...

Inter-laboratory comparisons of hexenuronic acid measurements in kraft eucalyptus pulps using a UV-Vis spectroscopic method

Treesearch

J.Y. Zhu; H.F Zhou; Chai X.S.; Donna Johannes; Richard Pope; Cristina Valls; M. Blanca Roncero

2014-01-01

An inter-laboratory comparison of a UV-Vis spectroscopic method (TAPPI T 282 om-13 “Hexeneuronic acid content of chemical pulp”) for hexeneuronic acid measurements was conducted using three eucalyptus kraft pulps. The pulp samples were produced in a laboratory at kappa numbers of approximately 14, 20, and 35. The hexeneuronic acid contents of the three pulps were...

Inter-laboratory comparison of the in vivo comet assay including three image analysis systems.

PubMed

Plappert-Helbig, Ulla; Guérard, Melanie

2015-12-01

To compare the extent of potential inter-laboratory variability and the influence of different comet image analysis systems, in vivo comet experiments were conducted using the genotoxicants ethyl methanesulfonate and methyl methanesulfonate. Tissue samples from the same animals were processed and analyzed-including independent slide evaluation by image analysis-in two laboratories with extensive experience in performing the comet assay. The analysis revealed low inter-laboratory experimental variability. Neither the use of different image analysis systems, nor the staining procedure of DNA (propidium iodide vs. SYBR® Gold), considerably impacted the results or sensitivity of the assay. In addition, relatively high stability of the staining intensity of propidium iodide-stained slides was found in slides that were refrigerated for over 3 months. In conclusion, following a thoroughly defined protocol and standardized routine procedures ensures that the comet assay is robust and generates comparable results between different laboratories. © 2015 Wiley Periodicals, Inc.

POTENTIAL RADIOACTIVE POLLUTANTS RESULTING FROM EXPANDED ENERGY PROGRAMS

EPA Science Inventory

An effective environmental monitoring program must have a quality assurance component to assure the production of valid data. Quality assurance has many components: calibration standards, standard reference materials, standard reference methods, interlaboratory comparison studies...

Inter-laboratory analysis of selected genetically modified plant reference materials with digital PCR.

PubMed

Dobnik, David; Demšar, Tina; Huber, Ingrid; Gerdes, Lars; Broeders, Sylvia; Roosens, Nancy; Debode, Frederic; Berben, Gilbert; Žel, Jana

2018-01-01

Digital PCR (dPCR), as a new technology in the field of genetically modified (GM) organism (GMO) testing, enables determination of absolute target copy numbers. The purpose of our study was to test the transferability of methods designed for quantitative PCR (qPCR) to dPCR and to carry out an inter-laboratory comparison of the performance of two different dPCR platforms when determining the absolute GM copy numbers and GM copy number ratio in reference materials certified for GM content in mass fraction. Overall results in terms of measured GM% were within acceptable variation limits for both tested dPCR systems. However, the determined absolute copy numbers for individual genes or events showed higher variability between laboratories in one third of the cases, most possibly due to variability in the technical work, droplet size variability, and analysis of the raw data. GMO quantification with dPCR and qPCR was comparable. As methods originally designed for qPCR performed well in dPCR systems, already validated qPCR assays can most generally be used for dPCR technology with the purpose of GMO detection. Graphical abstract The output of three different PCR-based platforms was assessed in an inter-laboratory comparison.

Inter-laboratory comparison of X-ray fluorescence analyses of eruptive products of El Chichón Volcano, Chiapas, Mexico

USGS Publications Warehouse

Tilling, Robert I.; Bornhorst, Theodore J.; Taggart, Joseph E.; Rose, William I.; McGee, James J.

1987-01-01

An inter-laboratory comparison has been made of X-ray fluorescence analyses of 10 samples of lava and pumices from El Chichón Volcano, Chiapas, Mexico. Some determinations of major-element constituents agree within analytical uncertainty, whereas others exchibit significant bias. Analyses carried out at the Michigan Technological University (MTU) laboratory are systematically lower in MgO (26–48%), Fetotal(5–18%), CaO (4–15%) and higher in K2O (0–15%) than analyses made at the U.S. Geological Survey (USGS) Denver laboratory. These differences are ascribed in part to a complex combination of calibration assumptionsand mineralogical and particle-size effects inherent in the use of pressed rock-powder pellets in the analytical procedure of the MTU laboratory. Other, but as yet unknown, differences in sample preparation and/or analytical technique may also be important; effects related to natural sample inhomogeneityare believed to be insignificant. The inter-laboratory differences in the analytical data complicated accurate assessment of whether El Chichón magmas have changed composition during the past 300 000 a. Knowledge of such change is needed for understanding petrogenetic history and for such related studies as evaluation of volcanic hazards.

Global Inter-Laboratory Fecal Source Identification Methods Comparison Study

EPA Science Inventory

Source tracking is key to identifying sources of fecal contamination for remediation as well as risk assessment. Previous intra- and inter-lab studies have investigated the performance of human and cow-associated source tracking markers, as well as library-dependent fecal source ...

77 FR 29757 - Guidelines Establishing Test Procedures for the Analysis of Pollutants Under the Clean Water Act...

Federal Register 2010, 2011, 2012, 2013, 2014

2012-05-18

... Discharge Elimination System (NPDES) program, issue permits with conditions designed to ensure compliance... completion of a full inter-laboratory validation study designed to fully characterize the performance of...

[Inter-laboratory reproducibility of pulsed-field electrophoresis for the study of 12 types of Pseudomonas aeruginosa].

PubMed

Foissaud, V; Puyhardy, J M; Chapalain, J C; Salord, H; Depina, J J; Morillon, M; Nicolas, P; Perrier-Gros-Claude, J D

1999-12-01

The increasing hospital-to-hospital transmission of multiple drug-resistant bacteria is a major concern for bacteriology laboratories involved in nosocomial infection control. The interlaboratory reproducibility of pulsed-field gel electrophoresis (PFGE) for Pseudomonas aeruginosa typing was evaluated by asking four hospital laboratories (two in Lyon, one in Brest, and one in Marseille) to study 11 P. aeruginosa isolates, some of which were epidemiologically related, and the reference strain ATCC 27853. Two laboratories used the Genepath system, one the Chef DR II, system, and one the Chef Mapper system, Bio-Rad, restriction/Spe I. Profiles were read visually and by computerized comparison of restriction band molecular weights (Taxotron, software, PAD Grimont, Pasteur Institute, Paris, France). These two methods led to similar epidemiological conclusions. However, centralization of the data showed poor center-to-center reproducibility due to inadequate standardization of the procedure.

Interlaboratory study for nickel alloy 625 made by laser powder bed fusion to quantify mechanical property variability.

PubMed

Brown, Christopher U; Jacob, Gregor; Stoudt, Mark; Moylan, Shawn; Slotwinski, John; Donmez, Alkan

2016-08-01

Six different organizations participated in this interlaboratory study to quantify the variability in the tensile properties of Inconel 625 specimens manufactured using laser-powder-bed-fusion additive manufacturing machines. The tensile specimens were heat treated and tensile tests conducted until failure. The properties measured were yield strength, ultimate tensile strength, elastic modulus, and elongation. Statistical analysis revealed that between-participant variability for yield strength, ultimate tensile strength, and elastic modulus values were significantly higher (up to 4 times) than typical within-participant variations. Only between-participant and within-participant variability were both similar for elongation. A scanning electron microscope was used to examine one tensile specimen for fractography. The fracture surface does not have many secondary cracks or other features that would reduce the mechanical properties. In fact, the features largely consist of microvoid coalescence and are entirely consistent with ductile failure.

Interlaboratory study for nickel alloy 625 made by laser powder bed fusion to quantify mechanical property variability

PubMed Central

Brown, Christopher U.; Jacob, Gregor; Stoudt, Mark; Moylan, Shawn; Slotwinski, John; Donmez, Alkan

2017-01-01

Six different organizations participated in this interlaboratory study to quantify the variability in the tensile properties of Inconel 625 specimens manufactured using laser-powder-bed-fusion additive manufacturing machines. The tensile specimens were heat treated and tensile tests conducted until failure. The properties measured were yield strength, ultimate tensile strength, elastic modulus, and elongation. Statistical analysis revealed that between-participant variability for yield strength, ultimate tensile strength, and elastic modulus values were significantly higher (up to 4 times) than typical within-participant variations. Only between-participant and within-participant variability were both similar for elongation. A scanning electron microscope was used to examine one tensile specimen for fractography. The fracture surface does not have many secondary cracks or other features that would reduce the mechanical properties. In fact, the features largely consist of microvoid coalescence and are entirely consistent with ductile failure. PMID:28243032

Interlaboratory Study for Nickel Alloy 625 Made by Laser Powder Bed Fusion to Quantify Mechanical Property Variability

NASA Astrophysics Data System (ADS)

Brown, Christopher U.; Jacob, Gregor; Stoudt, Mark; Moylan, Shawn; Slotwinski, John; Donmez, Alkan

2016-08-01

Six different organizations participated in this interlaboratory study to quantify the variability in the tensile properties of Inconel 625 specimens manufactured using laser powder bed fusion-additive manufacturing machines. The tensile specimens were heat treated and tensile tests were conducted until failure. The properties measured were yield strength, ultimate tensile strength, elastic modulus, and elongation. Statistical analysis revealed that between-participant variability for yield strength, ultimate tensile strength, and elastic modulus values were significantly higher (up to four times) than typical within-participant variations. Only between-participant and within-participant variability were both similar for elongation. A scanning electron microscope was used to examine one tensile specimen for fractography. The fracture surface does not have many secondary cracks or other features that would reduce the mechanical properties. In fact, the features largely consist of microvoid coalescence and are entirely consistent with ductile failure.

Interlaboratory Comparison of Methods Determining the Botanical Composition of Animal Feed.

PubMed

Braglia, Luca; Morello, Laura; Gavazzi, Floriana; Gianì, Silvia; Mastromauro, Francesco; Breviario, Diego; Cardoso, Hélia Guerra; Valadas, Vera; Campos, Maria Doroteia

2018-01-01

A consortium of European enterprises and research institutions has been engaged in the Feed-Code Project with the aim of addressing the requirements stated in European Union Regulation No. 767/2009, concerning market placement and use of feed of known and ascertained botanical composition. Accordingly, an interlaboratory trial was set up to compare the performance of different assays based either on optical microscope or DNA analysis for the qualitative and quantitative identification of the composition of compound animal feeds. A tubulin-based polymorphism method, on which the Feed-Code platform was developed, provided the most accurate results. The present study highlights the need for the performance of ring trials for the determination of the botanical composition of animal feeds and raises an alarm on the actual status of analytical inaccuracy.

Interlaboratory Comparison of Magnetic Thin Film Measurements.

PubMed

da Silva, F C S; Wang, C M; Pappas, D P

2003-01-01

A potential low magnetic moment standard reference material (SRM) was studied in an interlaboratory comparison. The mean and the standard deviation of the saturation moment m s, the remanent moment m r, and the intrinsic coercivity H c of nine samples were extracted from hysteresis-loop measurements. Samples were measured by thirteen laboratories using inductive-field loopers, vibrating-sample magnetometers, alternating-gradient force magnetometers, and superconducting quantum-interference-device magnetometers. NiFe films on Si substrates had saturation moment measurements reproduced within 5 % variation among the laboratories. The results show that a good candidate for an SRM must have a highly square hysteresis loop (m r/m s > 90 %), H c ≈ 400 A·m(-1) (5 Oe), and m s ≈ 2 × 10(-7) A·m(2) (2 × 10(-4) emu).

Interlaboratory comparability of serum cotinine measurements at smoker and nonsmoker concentration levels: A round-robin study

PubMed Central

Jacob, Peyton; Holiday, David B.; Benowitz, Neal L.; Sosnoff, Connie S.; Doig, Mira V.; Feyerabend, Colin; Aldous, Kenneth M.; Sharifi, Mehran; Kellogg, Mark D.; Langman, Loralie J.

2009-01-01

Introduction: Cotinine, the primary proximate metabolite of nicotine, is commonly measured as an index of exposure to tobacco in both active users of tobacco and nonsmokers with possible exposure to secondhand smoke (SHS). A number of laboratories have implemented analyses for measuring serum cotinine in recent years, but there have been few interlaboratory comparisons of the results. Among nonsmokers exposed to SHS, the concentration of cotinine in blood can be quite low, and extensive variability in these measurements has been reported in the past. Methods: In this study, a group of seven laboratories, all experienced in serum cotinine analysis, measured eight coded serum pools with concentrations ranging from background levels of about 0.05 ng/ml to relatively high concentrations in the active smokers range. All laboratories used either gas–liquid chromatography with nitrogen–phosphorus detection or liquid chromatography with mass spectrometric detection. Results: All seven laboratories reliably measured the cotinine concentrations in samples that were within the range of their methods. In each case, the results for the pools were correctly ranked in order, and no significant interlaboratory bias was observed at the 5% level of significance for results from any of the pools. Discussion: We conclude that present methods of chromatographic analysis of serum cotinine, as used by these experienced laboratories, are capable of providing accurate and precise results in both the smoker and the nonsmoker concentration range. PMID:19933777

Amino acid racemization dating of fossil bones, I. inter-laboratory comparison of racemization measurements

USGS Publications Warehouse

Bada, J.L.; Hoopes, E.; Darling, D.; Dungworth, G.; Kessels, H.J.; Kvenvolden, K.A.; Blunt, D.J.

1979-01-01

Enantiomeric measurements for aspartic acid, glutamic acid, and alanine in twenty-one different fossil bone samples have been carried out by three different laboratories using different analytical methods. These inter-laboratory comparisons demonstrate that D/L aspartic acid measurements are highly reproducible, whereas the enantiomeric measurements for the other amino acids show a wide variation between the three laboratories. At present, aspartic acid measurements are the most suitable for racemization dating of bone because of their superior analytical precision. ?? 1979.

Use of Web-based training for quality improvement between a field immunohistochemistry laboratory in Nigeria and its United States-based partner institution.

PubMed

Oluwasola, Abideen O; Malaka, David; Khramtsov, Andrey Ilyich; Ikpatt, Offiong Francis; Odetunde, Abayomi; Adeyanju, Oyinlolu Olorunsogo; Sveen, Walmy Elisabeth; Falusi, Adeyinka Gloria; Huo, Dezheng; Olopade, Olufunmilayo Ibironke

2013-12-01

The importance of hormone receptor status in assigning treatment and the potential use of human epidermal growth factor receptor 2 (HER2)-targeted therapy have made it beneficial for laboratories to improve detection techniques. Because interlaboratory variability in immunohistochemistry (IHC) tests may also affect studies of breast cancer subtypes in different countries, we undertook a Web-based quality improvement training and a comparative study of accuracy of immunohistochemical tests of breast cancer biomarkers between a well-established laboratory in the United States (University of Chicago) and a field laboratory in Ibadan, Nigeria. Two hundred and thirty-two breast tumor blocks were evaluated for estrogen receptors (ERs), progesterone receptors (PRs), and HER2 status at both laboratories using tissue microarray technique. Initially, concordance analysis revealed κ scores of 0.42 (moderate agreement) for ER, 0.41 (moderate agreement) for PR, and 0.39 (fair agreement) for HER2 between the 2 laboratories. Antigen retrieval techniques and scoring methods were identified as important reasons for discrepancy. Web-based conferences using Web conferencing tools such as Skype and WebEx were then held periodically to discuss IHC staining protocols and standard scoring systems and to resolve discrepant cases. After quality assurance and training, the agreement improved to 0.64 (substantial agreement) for ER, 0.60 (moderate agreement) for PR, and 0.75 (substantial agreement) for HER2. We found Web-based conferences and digital microscopy useful and cost-effective tools for quality assurance of IHC, consultation, and collaboration between distant laboratories. Quality improvement exercises in testing of tumor biomarkers will reduce misclassification in epidemiologic studies of breast cancer subtypes and provide much needed capacity building in resource-poor countries. © 2013.

Results of an inter-laboratory study of glass formulation for the immobilization of excess plutonium

DOE Office of Scientific and Technical Information (OSTI.GOV)

Peeler, D.K.

1999-12-08

The primary focus of the current study is to determine allowable loadings of feed streams containing different ratios of plutonium, uranium, and minor components into the LaBS glass and to evaluate thermal stability with respect to the DWPF pour.

Intact cell mass spectrometry (ICMS) used to type methicillin-resistant Staphylococcus aureus: media effects and inter-laboratory reproducibility.

PubMed

Walker, J; Fox, A J; Edwards-Jones, V; Gordon, D B

2002-02-01

Intact cell mass spectrometry (ICMS) rapidly analyses the surface composition of microorganisms providing rapid, discriminatory fingerprints for identification and subtyping of important nosocomial pathogens such as methicillin resistant Staphylocccus aureus (MRSA). In this study, ICMS using matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI TOF/MS) was assessed for the identification and subtyping of MRSA. An intra- and inter-laboratory reproducibility study was carried out and the effects of culture media (an important source of variation for ICMS) were also studied. Several media used for the cultural identification of MRSA were examined using a panel of well-characterised staphylococcal isolates (n=26). Six MRSA isolates were analysed over a 1-month period for intra-laboratory reproducibility on the same instrument and three different culture media. Spectra were consistent for each isolate between the four experiments on the same culture medium. Individual isolates produced different spectral profiles on different culture media. Spectra from organisms grown on Columbia blood agar contained more peaks (approximately 120) compared to Columbia agar (approximately 50) and methicillin mannitol salt agar (approximately 25). All 26 staphylococcal isolates were subjected to an inter-laboratory study on two MALDI instruments. For each isolate, the overall spectral profile was the same for each of the two instruments but the baseline threshold values was adjusted due to instrument differences in detector sensitivities. Differences between certain regions of the spectra reproducibly identified isolates belonging to the two major MRSA strains (EMRSA phage group 15 and 16). These results demonstrate ICMS with appropriate media selection is a rapid and reproducible technique for identification and discrimination of MRSA.

Analytical Round Robin for Elastic-Plastic Analysis of Surface Cracked Plates, Phase II Results

NASA Technical Reports Server (NTRS)

Allen, P. A.; Wells, D. N.

2017-01-01

The second phase of an analytical round robin for the elastic-plastic analysis of surface cracks in flat plates was conducted under the auspices of ASTM Interlaboratory Study 732. The interlaboratory study (ILS) had 10 participants with a broad range of expertise and experience, and experimental results from a surface crack tension test in 4142 steel plate loaded well into the elastic-plastic regime provided the basis for the study. The participants were asked to evaluate a surface crack tension test according to the version of the surface crack initiation toughness testing standard published at the time of the ILS, E2899-13. Data were provided to each participant that represent the fundamental information that would be provided by a mechanical test laboratory prior to evaluating the test result. Overall, the participant’s test analysis results were in good agreement and constructive feedback was received that has resulted in an improved published version of the standard E2899-15.

Advancing the Use of Passive Sampling in Risk Assessment and Management of Sediments Contaminated with Hydrophobic Organic Chemicals: Results of an International Ex Situ Passive Sampling Interlaboratory Comparison

PubMed Central

2018-01-01

This work presents the results of an international interlaboratory comparison on ex situ passive sampling in sediments. The main objectives were to map the state of the science in passively sampling sediments, identify sources of variability, provide recommendations and practical guidance for standardized passive sampling, and advance the use of passive sampling in regulatory decision making by increasing confidence in the use of the technique. The study was performed by a consortium of 11 laboratories and included experiments with 14 passive sampling formats on 3 sediments for 25 target chemicals (PAHs and PCBs). The resulting overall interlaboratory variability was large (a factor of ∼10), but standardization of methods halved this variability. The remaining variability was primarily due to factors not related to passive sampling itself, i.e., sediment heterogeneity and analytical chemistry. Excluding the latter source of variability, by performing all analyses in one laboratory, showed that passive sampling results can have a high precision and a very low intermethod variability (

Comparison of bulk sediment and sediment elutriate toxicity testing methods

EPA Science Inventory

Elutriate bioassays are among numerous methods that exist for assessing the potential toxicity of sediments in aquatic systems. In this study, interlaboratory results were compared from 96-hour Ceriodaphnia dubia and Pimephales promelas static-renewal acute toxicity tests conduct...

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lindahl, P.C.

A proposed American Society for Testing and Materials (ASTM) method for the determination of arsenic and selenium content in coal has been used and evaluated in the Analytical Chemistry Laboratory (ACL) at Argonne National Laboratory (ANL) as part of an interlaboratory study. Coal is conducted with Eschka's mixture (MgO + Na/sub 2/CO/sub 3/), followed by determination of the aresnic and selenium content by hydride generation/atomic absorption spectrophotometry. The method was evaluated on a series of coals, including two National Bureau of Standards-Standards Reference Material (NBS-SRM) coals and twelve ASTM coal samples. Comparison of ACL/ANL arsenic and selenium data for themore » suite of coal analyzed showed excellent agreement with certified values for the NBS-SRM coals and with interlaboratory data from five other laboratories for the ASTM coals. 11 refs., 3 figs., 6 tabs.« less

An inter-laboratory comparison study of the ANSI/BIFMA standard test method M7.1 for furniture

EPA Science Inventory

Five laboratories using five different test chambers participated in the study to quantify within- and between-laboratory variability in the measurement of emissions of volatile organic compounds (VOCs) from new commercial furniture test items following ANSI/BIFMA M7.1. Test item...

INTERLABORATORY STUDY OF THE COLD VAPOR TECHNIQUE FOR TOTAL MERCURY IN WATER

EPA Science Inventory

The American Society for Testing and Materials (ASTM) and the U.S. Environmental Protection Agency (EPA) conducted a joint study of the cold vapor technique for total mercury in water, before formal acceptance of the method by each organization. The method employs an acid-permang...

INTERLABORATORY COMPARISON STUDIES FOR CHARACTERIZATION OF ORGANIC COMPOUNDS IN PARTICULATE MATTER

EPA Science Inventory

A working group of investigators, who are characterizing and quantifying the organic compounds in particulate matter (PM) as part of the US EPA's PM 2.5 research program and related studies, was established three years ago to advance the quality and comparability of data on the...

Practical methodological guide for hydrometric inter-laboratory organisation

NASA Astrophysics Data System (ADS)

Besson, David; Bertrand, Xavier

2015-04-01

Discharge measurements performed by the French governmental hydrometer team feed a national database. This data is available for general river flows knowkedge, flood forecasting, low water survey, statistical calculations flow, control flow regulatory and many other uses. Regularly checking the measurements quality and better quantifying its accuracy is therefore an absolute need. The practice of inter-laboratory comparison in hydrometry particularly developed during the last decade. Indeed, discharge measurement can not easily be linked to a standard. Therefore, on-site measurement accuracy control is very difficult. Inter-laboratory comparison is thus a practical solution to this issue. However, it needs some regulations in order to ease its practice and legitimize its results. To do so, the French government hydrometrics teams produced a practical methodological guide for hydrometric inter-laboratory organisation in destination of hydrometers community in view of ensure the harmonization of inter-laboratory comparison practices for different materials (ADCP, current meter on wadind rod or gauging van, tracer dilution, surface speed) and flow range (flood, low water). Ensure the results formalization and banking. The realisation of this practice guide is grounded on the experience of the governmental teams & their partners (or fellows), following existing approaches (Doppler group especially). The guide is designated to validate compliance measures and identify outliers : Hardware, methodological, environmental, or human. Inter-laboratory comparison provides the means to verify the compliance of the instruments (devices + methods + operators) and provides methods to determine an experimental uncertainty of the tested measurement method which is valid only for the site and the measurement conditions but does not address the calibration or periodic monitoring of the few materials. After some conceptual definitions, the guide describes the different stages of an inter-comparison campaign: the campaing creation: targets, participants ( instruments type and number) and site preparation of test protocols and schedule; the campaign set-up (organization): invitation and pre-information of the participants, logistics, field preparation; the campaign conduct: participants reception and information, sequences of tests, results analysis and communication, balance sheet; post-campaign work: further analysis, dissemination and periodic verification of the instruments. This guide is associated with measurement instruments forms, reminding their limits and conditions for use, land forms, used to record all the necessary information during the inter-comparison campaign (site description and measurement conditions, equipment and its settings, and the set of measurements or intermediate calculations to the final results) as well as a calculation tool and banking measures and results.

MGDB: crossing the marker genes of a user microarray with a database of public-microarrays marker genes.

PubMed

Huerta, Mario; Munyi, Marc; Expósito, David; Querol, Enric; Cedano, Juan

2014-06-15

The microarrays performed by scientific teams grow exponentially. These microarray data could be useful for researchers around the world, but unfortunately they are underused. To fully exploit these data, it is necessary (i) to extract these data from a repository of the high-throughput gene expression data like Gene Expression Omnibus (GEO) and (ii) to make the data from different microarrays comparable with tools easy to use for scientists. We have developed these two solutions in our server, implementing a database of microarray marker genes (Marker Genes Data Base). This database contains the marker genes of all GEO microarray datasets and it is updated monthly with the new microarrays from GEO. Thus, researchers can see whether the marker genes of their microarray are marker genes in other microarrays in the database, expanding the analysis of their microarray to the rest of the public microarrays. This solution helps not only to corroborate the conclusions regarding a researcher's microarray but also to identify the phenotype of different subsets of individuals under investigation, to frame the results with microarray experiments from other species, pathologies or tissues, to search for drugs that promote the transition between the studied phenotypes, to detect undesirable side effects of the treatment applied, etc. Thus, the researcher can quickly add relevant information to his/her studies from all of the previous analyses performed in other studies as long as they have been deposited in public repositories. Marker-gene database tool: http://ibb.uab.es/mgdb © The Author 2014. Published by Oxford University Press.

Interlaboratory validation of the modified murine local lymph node assay based on adenosine triphosphate measurement.

PubMed

Omori, Takashi; Idehara, Kenji; Kojima, Hajime; Sozu, Takashi; Arima, Kazunori; Goto, Hirohiko; Hanada, Tomohiko; Ikarashi, Yoshiaki; Inoda, Taketo; Kanazawa, Yukiko; Kosaka, Tadashi; Maki, Eiji; Morimoto, Takashi; Shinoda, Shinsuke; Shinoda, Naoki; Takeyoshi, Masahiro; Tanaka, Masashi; Uratani, Mamoru; Usami, Masahito; Yamanaka, Atsushi; Yoneda, Tomofumi; Yoshimura, Isao; Yuasa, Atsuko

2008-01-01

The murine local lymph node assay (LLNA) is a well-established alternative to the guinea pig maximization test (GPMT) or Buehler test (BT) for the assessment of the skin sensitizing ability of drugs and chemicals. Daicel Chemical Industries Ltd. has developed a modified LLNA based on the adenosine triphosphate (ATP) content (LLNA-DA). We conducted 2 interlaboratory validation studies to evaluate the reliability and relevance of LLNA-DA. The experiment involved 17 laboratories, wherein 14 chemicals were examined under blinded conditions. In the first study, 3 chemicals were examined in 10 laboratories and the remaining 9 were examined in 3 laboratories. In the second study, 1 chemical was examined in 7 laboratories and the remaining 4 chemicals were examined in 4 laboratories. The data were expressed as the ATP content for each chemical-treated group, and the stimulation index (SI) for each chemical-treated group was determined as the increase in the ATP content relative to the concurrent vehicle control group. An SI of 3 was set as the cut-off value for exhibiting skin sensitization activity. The results of the first study obtained in the experiments conducted for the 3 chemicals that were examined in all the 10 laboratories and for 5 of the remaining 9 chemicals were sufficiently consistent with small variations in their SI values. The sensitivity, specificity, and accuracy of LLNA-DA against those of GPMT/BT were 7/8 (87.5%), 3/3 (100%), and 10/11 (90.9%), respectively. In the second study, all the 5 chemicals studied demonstrated acceptably small interlaboratory variations. In the first study, a large variation was observed for 2 chemicals; in the second study, this variation was small. It was attributed to the application of dimethylsulfoxide as the solvent for the metallic salts. In conclusion, these 2 studies provide good evidence for the reliability of the LLNA-DA.

Interlaboratory comparison of δ13C and δD measurements of atmospheric CH4 for combined use of data sets from different laboratories

NASA Astrophysics Data System (ADS)

Umezawa, Taku; Brenninkmeijer, Carl A. M.; Röckmann, Thomas; van der Veen, Carina; Tyler, Stanley C.; Fujita, Ryo; Morimoto, Shinji; Aoki, Shuji; Sowers, Todd; Schmitt, Jochen; Bock, Michael; Beck, Jonas; Fischer, Hubertus; Michel, Sylvia E.; Vaughn, Bruce H.; Miller, John B.; White, James W. C.; Brailsford, Gordon; Schaefer, Hinrich; Sperlich, Peter; Brand, Willi A.; Rothe, Michael; Blunier, Thomas; Lowry, David; Fisher, Rebecca E.; Nisbet, Euan G.; Rice, Andrew L.; Bergamaschi, Peter; Veidt, Cordelia; Levin, Ingeborg

2018-03-01

We report results from a worldwide interlaboratory comparison of samples among laboratories that measure (or measured) stable carbon and hydrogen isotope ratios of atmospheric CH4 (δ13C-CH4 and δD-CH4). The offsets among the laboratories are larger than the measurement reproducibility of individual laboratories. To disentangle plausible measurement offsets, we evaluated and critically assessed a large number of intercomparison results, some of which have been documented previously in the literature. The results indicate significant offsets of δ13C-CH4 and δD-CH4 measurements among data sets reported from different laboratories; the differences among laboratories at modern atmospheric CH4 level spread over ranges of 0.5 ‰ for δ13C-CH4 and 13 ‰ for δD-CH4. The intercomparison results summarized in this study may be of help in future attempts to harmonize δ13C-CH4 and δD-CH4 data sets from different laboratories in order to jointly incorporate them into modelling studies. However, establishing a merged data set, which includes δ13C-CH4 and δD-CH4 data from multiple laboratories with desirable compatibility, is still challenging due to differences among laboratories in instrument settings, correction methods, traceability to reference materials and long-term data management. Further efforts are needed to identify causes of the interlaboratory measurement offsets and to decrease those to move towards the best use of available δ13C-CH4 and δD-CH4 data sets.

Reliability on intra-laboratory and inter-laboratory data of hair mineral analysis comparing with blood analysis.

PubMed

Namkoong, Sun; Hong, Seung Phil; Kim, Myung Hwa; Park, Byung Cheol

2013-02-01

Nowadays, although its clinical value remains controversial institutions utilize hair mineral analysis. Arguments about the reliability of hair mineral analysis persist, and there have been evaluations of commercial laboratories performing hair mineral analysis. The objective of this study was to assess the reliability of intra-laboratory and inter-laboratory data at three commercial laboratories conducting hair mineral analysis, compared to serum mineral analysis. Two divided hair samples taken from near the scalp were submitted for analysis at the same time, to all laboratories, from one healthy volunteer. Each laboratory sent a report consisting of quantitative results and their interpretation of health implications. Differences among intra-laboratory and interlaboratory data were analyzed using SPSS version 12.0 (SPSS Inc., USA). All the laboratories used identical methods for quantitative analysis, and they generated consistent numerical results according to Friedman analysis of variance. However, the normal reference ranges of each laboratory varied. As such, each laboratory interpreted the patient's health differently. On intra-laboratory data, Wilcoxon analysis suggested they generated relatively coherent data, but laboratory B could not in one element, so its reliability was doubtful. In comparison with the blood test, laboratory C generated identical results, but not laboratory A and B. Hair mineral analysis has its limitations, considering the reliability of inter and intra laboratory analysis comparing with blood analysis. As such, clinicians should be cautious when applying hair mineral analysis as an ancillary tool. Each laboratory included in this study requires continuous refinement from now on for inducing standardized normal reference levels.

Comparison of Different Matrices as Potential Quality Control Samples for Neurochemical Dementia Diagnostics.

PubMed

Lelental, Natalia; Brandner, Sebastian; Kofanova, Olga; Blennow, Kaj; Zetterberg, Henrik; Andreasson, Ulf; Engelborghs, Sebastiaan; Mroczko, Barbara; Gabryelewicz, Tomasz; Teunissen, Charlotte; Mollenhauer, Brit; Parnetti, Lucilla; Chiasserini, Davide; Molinuevo, Jose Luis; Perret-Liaudet, Armand; Verbeek, Marcel M; Andreasen, Niels; Brosseron, Frederic; Bahl, Justyna M C; Herukka, Sanna-Kaisa; Hausner, Lucrezia; Frölich, Lutz; Labonte, Anne; Poirier, Judes; Miller, Anne-Marie; Zilka, Norbert; Kovacech, Branislav; Urbani, Andrea; Suardi, Silvia; Oliveira, Catarina; Baldeiras, Ines; Dubois, Bruno; Rot, Uros; Lehmann, Sylvain; Skinningsrud, Anders; Betsou, Fay; Wiltfang, Jens; Gkatzima, Olymbia; Winblad, Bengt; Buchfelder, Michael; Kornhuber, Johannes; Lewczuk, Piotr

2016-03-01

Assay-vendor independent quality control (QC) samples for neurochemical dementia diagnostics (NDD) biomarkers are so far commercially unavailable. This requires that NDD laboratories prepare their own QC samples, for example by pooling leftover cerebrospinal fluid (CSF) samples. To prepare and test alternative matrices for QC samples that could facilitate intra- and inter-laboratory QC of the NDD biomarkers. Three matrices were validated in this study: (A) human pooled CSF, (B) Aβ peptides spiked into human prediluted plasma, and (C) Aβ peptides spiked into solution of bovine serum albumin in phosphate-buffered saline. All matrices were tested also after supplementation with an antibacterial agent (sodium azide). We analyzed short- and long-term stability of the biomarkers with ELISA and chemiluminescence (Fujirebio Europe, MSD, IBL International), and performed an inter-laboratory variability study. NDD biomarkers turned out to be stable in almost all samples stored at the tested conditions for up to 14 days as well as in samples stored deep-frozen (at - 80°C) for up to one year. Sodium azide did not influence biomarker stability. Inter-center variability of the samples sent at room temperature (pooled CSF, freeze-dried CSF, and four artificial matrices) was comparable to the results obtained on deep-frozen samples in other large-scale projects. Our results suggest that it is possible to replace self-made, CSF-based QC samples with large-scale volumes of QC materials prepared with artificial peptides and matrices. This would greatly facilitate intra- and inter-laboratory QC schedules for NDD measurements.

Contributions to Statistical Problems Related to Microarray Data

ERIC Educational Resources Information Center

Hong, Feng

2009-01-01

Microarray is a high throughput technology to measure the gene expression. Analysis of microarray data brings many interesting and challenging problems. This thesis consists three studies related to microarray data. First, we propose a Bayesian model for microarray data and use Bayes Factors to identify differentially expressed genes. Second, we…

Use of the rapid immersion test to evaluate the efficacy of admixtures to mitigate alkali-silica reactivity.

DOT National Transportation Integrated Search

1996-01-01

The Federal Highway Administration (FHWA) is conducting an interlaboratory study to validate certain findings in the Strategic Highway Research Program (SHRP) C-343 report on alkali-silica reactivity. This paper reports the Virginia Transportation Re...

Collaborative analysis of wheat endosperm compressive material properties

USDA-ARS?s Scientific Manuscript database

The objective measurement of cereal endosperm texture, for wheat (Triticum L.) in particular, is relevant to the milling, processing and utilization of grain. The objective of this study was to evaluate the inter-laboratory results of compression failure testing of wheat endosperm specimens of defi...

An Experimental Introduction to Interlaboratory Exercises in Analytical Chemistry

ERIC Educational Resources Information Center

Puignou, L.; Llaurado, M.

2005-01-01

An experimental exercise on analytical proficiency studies in collaborative trials is proposed. This practical provides students in advanced undergraduate courses in chemistry, pharmacy, and biochemistry, with the opportunity to improve their quality assurance skills. It involves an environmental analysis, determining the concentration of a…

Fish species identification using PCR-RFLP analysis and lab-on-a-chip capillary electrophoresis: application to detect white fish species in food products and an interlaboratory study.

PubMed

Dooley, John J; Sage, Helen D; Clarke, Marie-Anne L; Brown, Helen M; Garrett, Stephen D

2005-05-04

Identification of 10 white fish species associated with U.K. food products was achieved using PCR-RFLP of the mitochondrial cytochrome b gene. Use of lab-on-a-chip capillary electrophoresis for end-point analysis enabled accurate sizing of DNA fragments and identification of fish species at a level of 5% (w/w) in a fish admixture. One restriction enzyme, DdeI, allowed discrimination of eight species. When combined with NlaIII and HaeIII, specific profiles for all 10 species were generated. The method was applied to a range of products and subjected to an interlaboratory study carried out by five U.K. food control laboratories. One hundred percent correct identification of single species samples and six of nine admixture samples was achieved by all laboratories. The results indicated that fish species identification could be carried out using a database of PCR-RFLP profiles without the need for reference materials.

Liquid chromatographic determination of histamine in fish, sauerkraut, and wine: interlaboratory study.

PubMed

Beljaars, P R; Van Dijk, R; Jonker, K M; Schout, L J

1998-01-01

An interlaboratory study of the liquid chromatographic (LC) determination of histamine in fish, sauerkraut, and wine was conducted. Diminuted and homogenized samples were suspended in water followed by clarification of extracts with perchloric acid, filtration, and dilution with water. After LC separation on a reversed-phase C18 column with phosphate buffer (pH 3.0)--acetonitrile (875 + 125, v/v) as mobile phase, histamine was measured fluorometrically (excitation, 340 nm; emission, 455 nm) in samples and standards after postcolumn derivatization with o-phthaldialdehyde (OPA). Fourteen samples (including 6 blind duplicates and 1 split level) containing histamine at about 10-400 mg/kg or mg/L were analyzed singly according to the proposed procedure by 11 laboratories. Results from one participant were excluded from statistical analysis. For all samples analyzed, repeatability relative standard deviations varied from 2.1 to 5.6%, and reproducibility relative standard deviations ranged from 2.2 to 7.1%. Averaged recoveries of histamine for this concentration range varied from 94 to 100%.

Degradation in PV Encapsulant Strength of Attachment: An Interlaboratory Study Towards a Climate-Specific Test

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miller, David C.; Annigoni, Eleonora; Ballion, Amal

Reduced strength of attachment of the encapsulant resulting from the outdoor environment, including ultraviolet (UV) radiation, may decrease photovoltaic (PV) module lifetime by enabling widespread corrosion of internal components. To date, few studies exist showing how the adhesion of PV components varies with environmental stress. We have conducted an interlaboratory experiment to provide an understanding that will be used to develop climatic specific module tests. Factors examined in the study included the UV light source (lamp type), temperature, and humidity to be proposed for use in accelerated aging tests. A poly (ethylene-co-vinyl acetate) (EVA) formulation often used in veteran PVmore » installations was studied using a compressive shear test - to quantify the strength of attachment at the EVA/glass interface. Replicate laminated glass/polymer/glass coupon specimens were weathered at 12 institutions using a variety of indoor chambers or field aging. Shear strength, shear strain, and toughness were measured using a mechanical load-frame for the compressive shear test, with subsequent optical imaging and electron microscopy of the separated surfaces.« less

Degradation in PV Encapsulation Strength of Attachment: An Interlaboratory Study Towards a Climate-Specific Test

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miller, David; Annigoni, Eleonora; Ballion, Amal

Reduced strength of attachment of the encapsulant resulting from the outdoor environment, including ultraviolet (UV) radiation, may decrease photovoltaic (PV) module lifetime by enabling widespread corrosion of internal components. To date, few studies exist showing how the adhesion of PV components varies with environmental stress. We have conducted an interlaboratory experiment to provide an understanding that will be used to develop climatic specific module tests. Factors examined in the study included the UV light source (lamp type), temperature, and humidity to be proposed for use in accelerated aging tests. A poly (ethylene-co-vinyl acetate) (EVA) formulation often used in veteran PVmore » installations was studied using a compressive shear test - to quantify the strength of attachment at the EVA/glass interface. Replicate laminated glass/polymer/glass coupon specimens were weathered at 12 institutions using a variety of indoor chambers or field aging. Shear strength, shear strain, and toughness were measured using a mechanical load-frame for the compressive shear test, with subsequent optical imaging and electron microscopy of the separated surfaces.« less

Degradation in PV Encapsulant Strength of Attachment: An Interlaboratory Study Towards a Climate-Specific Test: Preprint

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miller, David C.; Annigoni, Eleonora; Ballion, Amal

Reduced strength of attachment of the encapsulant resulting from outdoor environment, including ultraviolet (UV) radiation, may decrease photovoltaic (PV) module lifetime by enabling widespread corrosion of internal components. To date, few studies exist showing how the adhesion of PV components varies with environmental stress. We have conducted an interlaboratory experiment to provide an understanding that will be used to develop climatic specific module tests. Factors examined in the study included the UV light source (lamp type), temperature, and humidity to be proposed for use in accelerated aging tests. A poly (ethylene-co-vinyl acetate) (EVA) formulation often used in veteran PV installationsmore » was studied using a compressive shear test - to quantify the strength of attachment at the EVA/glass interface. Replicate laminated glass/polymer/glass coupon specimens were weathered at 12 institutions using a variety of indoor chambers or field aging. Shear strength, shear strain, and toughness were measured using a mechanical load-frame for the compressive shear test, with subsequent optical imaging and electron microscopy of the separated surfaces.« less

40 CFR Appendix D to Part 136 - Precision and Recovery Statements for Methods for Measuring Metals

Code of Federal Regulations, 2011 CFR

2011-07-01

... Accuracy Section with the following: Precision and Accuracy An interlaboratory study on metal analyses by... details are found in “USEPA Method Study 7, Analyses for Trace Methods in water by Atomic Absorption... study on metal analyses by this method was conducted by the Quality Assurance Branch (QAB) of the...

Performance Metrics for Liquid Chromatography-Tandem Mass Spectrometry Systems in Proteomics Analyses*

PubMed Central

Rudnick, Paul A.; Clauser, Karl R.; Kilpatrick, Lisa E.; Tchekhovskoi, Dmitrii V.; Neta, Pedatsur; Blonder, Nikša; Billheimer, Dean D.; Blackman, Ronald K.; Bunk, David M.; Cardasis, Helene L.; Ham, Amy-Joan L.; Jaffe, Jacob D.; Kinsinger, Christopher R.; Mesri, Mehdi; Neubert, Thomas A.; Schilling, Birgit; Tabb, David L.; Tegeler, Tony J.; Vega-Montoto, Lorenzo; Variyath, Asokan Mulayath; Wang, Mu; Wang, Pei; Whiteaker, Jeffrey R.; Zimmerman, Lisa J.; Carr, Steven A.; Fisher, Susan J.; Gibson, Bradford W.; Paulovich, Amanda G.; Regnier, Fred E.; Rodriguez, Henry; Spiegelman, Cliff; Tempst, Paul; Liebler, Daniel C.; Stein, Stephen E.

2010-01-01

A major unmet need in LC-MS/MS-based proteomics analyses is a set of tools for quantitative assessment of system performance and evaluation of technical variability. Here we describe 46 system performance metrics for monitoring chromatographic performance, electrospray source stability, MS1 and MS2 signals, dynamic sampling of ions for MS/MS, and peptide identification. Applied to data sets from replicate LC-MS/MS analyses, these metrics displayed consistent, reasonable responses to controlled perturbations. The metrics typically displayed variations less than 10% and thus can reveal even subtle differences in performance of system components. Analyses of data from interlaboratory studies conducted under a common standard operating procedure identified outlier data and provided clues to specific causes. Moreover, interlaboratory variation reflected by the metrics indicates which system components vary the most between laboratories. Application of these metrics enables rational, quantitative quality assessment for proteomics and other LC-MS/MS analytical applications. PMID:19837981

Interlaboratory Comparison Test as an Evaluation of Applicability of an Alternative Edible Oil Analysis by 1H NMR Spectroscopy.

PubMed

Zailer, Elina; Holzgrabe, Ulrike; Diehl, Bernd W K

2017-11-01

A proton (1H) NMR spectroscopic method was established for the quality assessment of vegetable oils. To date, several research studies have been published demonstrating the high potential of the NMR technique in lipid analysis. An interlaboratory comparison was organized with the following main objectives: (1) to evaluate an alternative analysis of edible oils by using 1H NMR spectroscopy; and (2) to determine the robustness and reproducibility of the method. Five different edible oil samples were analyzed by evaluating 15 signals (free fatty acids, peroxides, aldehydes, double bonds, and linoleic and linolenic acids) in each spectrum. A total of 21 NMR data sets were obtained from 17 international participant laboratories. The performance of each laboratory was assessed by their z-scores. The test was successfully passed by 90.5% of the participants. Results showed that NMR spectroscopy is a robust alternative method for edible oil analysis.

Inter-laboratory comparison of multi-locus variable-number tandem repeat analysis (MLVA) for verocytotoxin-producing Escherichia coli O157 to facilitate data sharing.

PubMed

Holmes, A; Perry, N; Willshaw, G; Hanson, M; Allison, L

2015-01-01

Multi-locus variable number tandem repeat analysis (MLVA) is used in clinical and reference laboratories for subtyping verocytotoxin-producing Escherichia coli O157 (VTEC O157). However, as yet there is no common allelic or profile nomenclature to enable laboratories to easily compare data. In this study, we carried out an inter-laboratory comparison of an eight-loci MLVA scheme using a set of 67 isolates of VTEC O157. We found all but two isolates were identical in profile in the two laboratories, and repeat units were homogeneous in size but some were incomplete. A subset of the isolates (n = 17) were sequenced to determine the actual copy number of representative alleles, thereby enabling alleles to be named according to international consensus guidelines. This work has enabled us to realize the potential of MLVA as a portable, highly discriminatory and convenient subtyping method.

Interlaboratory tests for low-level radioactivity measurements in France

DOE Office of Scientific and Technical Information (OSTI.GOV)

Despres, M.

In the framework of French national metrological coordination, the Laboratoire de Metrologie des Rayonnements Ionisants (LMRI) organizes every year an interlaboratory test program. The participation in these programs is on a voluntary basis and the results are presented in a nameless way. These tests concern radioactivity, neutron and dosimetry measurements. In this paper, a review of interlaboratory tests for low-level radioactivity measurements proposed by LMRI is given from 1975 to 1986. Every year, three types of tests are proposed: activity tests, multigamma tests, radioanalysis tests. For the last ten years, the number of results has been respectively 90 for activitymore » tests, 570 for multigamma tests and 400 for radioanalysis tests. The user's interest has increased, while the quality of results has improved. The deviations from LMRI's reference values are less than 3% for a restricted number of laboratories, and range between 3 and 10% for the majority of the others. The most urgent tests are for mixtures of {sup 90}Sr and activation and fission products in various matrices such as milk, grass, cereals, and sediments. 16 refs., 3 figs., 3 tabs.« less

Lupus anticoagulants: first French interlaboratory Etalonorme survey.

PubMed

Roussi, J; Roisin, J P; Goguel, A

1996-06-01

In 1994, the, French National Quality Control Group for Hematology, Etalonorme, conducted a large-scale interlaboratory survey concerning the detection of lupus anticoagulants (LA) involving all the 4,500 French laboratories. Each laboratory received the same batch of a lyophilized citrated plasma (94B3) prepared from a patient with LA that had been confirmed by all the techniques used in the intralaboratory study. In the interlaboratory survey, the screening test was activated partial thromboplastin time (APTT); mean APTT calculated from the results reported by 4,029 labs was prolonged (clotting ratio = 1.44) with a large dispersion (coefficients of variation = 18.8%). APTT of the mixture 94B3 + normal plasma were performed by 2,698 laboratories. No correction of APTT was obtained (R = 1.36, Rosner index = 24) with a wide variation between reagents (17 < Rosner index < 39). Only 15% of the participants performed confirmatory tests; dilute tissue thromboplastin inhibition test (TTI) performed by 509 laboratories gave 75% positive results. Tests with an increased amount of phospholipids (Staclot LA and Staclot PNP from Diagnostica Stago), used by 116 and 72 laboratories, gave 88% and 61% positive results, respectively. A total of 1,862 laboratories made the diagnosis of LA. The majority of those who failed in diagnosing LA used an APTT reagent largely used in France, containing kaolin. This survey allowed Etalonorme to inform French biologists and draft an educational program for the biologic detection of LA and the identification of its mechanism of action.

CAPILLARY GAS CHROMATOGRAPHY-ATOMIC EMISSION DETECTION METHOD FOR THE DETERMINATION OF PENTYLATED ORGANOTIN COMPOUNDS: INTERLABORATORY STUDY

EPA Science Inventory

A capillary gas chromatography-atomic emission detection (GC-AED) method was developed for the U. S. Environmental Protection Agency's Environmental Monitoring Systems Laboratory in Las Vegas, NV, for determination of selected organotin compounds. Here we report on an interlabora...

Interlaboratory comparison of reference materials for nitrogen-isotope-ratio measurements

USGS Publications Warehouse

Böhlke, John Karl; Coplen, Tyler B.

1995-01-01

Aliquots of seven different reference materials were distributed for an interlaboratory comparison of stable nitrogen-isotope-ratio measurements. Results from 15 laboratories were compiled and evaluated selectively to yield provisional values of 515N for each material, i, with respect to atmospheric N2 (o1SN,7air). The 515N values reported by the different laboratories are correlated in such a way that some of the major discrepancies may be removed by normalization (/. e., by altering the length of the ô N scale for each laboratory by an amount defined by local measurements of reference materials with extreme values).

Intra-Platform Repeatability and Inter-Platform Comparability of MicroRNA Microarray Technology

PubMed Central

Sato, Fumiaki; Tsuchiya, Soken; Terasawa, Kazuya; Tsujimoto, Gozoh

2009-01-01

Over the last decade, DNA microarray technology has provided a great contribution to the life sciences. The MicroArray Quality Control (MAQC) project demonstrated the way to analyze the expression microarray. Recently, microarray technology has been utilized to analyze a comprehensive microRNA expression profiling. Currently, several platforms of microRNA microarray chips are commercially available. Thus, we compared repeatability and comparability of five different microRNA microarray platforms (Agilent, Ambion, Exiqon, Invitrogen and Toray) using 309 microRNAs probes, and the Taqman microRNA system using 142 microRNA probes. This study demonstrated that microRNA microarray has high intra-platform repeatability and comparability to quantitative RT-PCR of microRNA. Among the five platforms, Agilent and Toray array showed relatively better performances than the others. However, the current lineup of commercially available microRNA microarray systems fails to show good inter-platform concordance, probably because of lack of an adequate normalization method and severe divergence in stringency of detection call criteria between different platforms. This study provided the basic information about the performance and the problems specific to the current microRNA microarray systems. PMID:19436744

Validation of standard method EN ISO 11290-part 2 for the enumeration of Listeria monocytogenes in food.

PubMed

Rollier, Patricia; Lombard, Bertrand; Guillier, Laurent; François, Danièle; Romero, Karol; Pierru, Sylvie; Bouhier, Laurence; Gnanou Besse, Nathalie

2018-05-01

The reference method for the detection and enumeration of L. monocytogenes in food (Standards EN ISO 11290-1&2) have been validated by inter-laboratory studies in the frame of the Mandate M381 from European Commission to CEN. In this paper, the inter-laboratory studies led in 2013 on 5 matrices (cold-smoked salmon, milk powdered infant food formula, vegetables, environment, and cheese) to validate Standard EN ISO 11290-2 are reported. According to the results obtained, the method of the revised Standard EN ISO 11290-2 can be considered as a good method for the enumeration of L. monocytogenes in foods and food processing environment, in particular for the matrices included in the study. Values of repeatability and reproducibility standard deviations can be considered satisfactory for this type of method with a confirmation stage, since most of them were below 0.3 log 10 , also at low levels, close to the regulatory limit of 100 CFU/g. Copyright © 2018 Elsevier B.V. All rights reserved.

Tools to minimize interlaboratory variability in vitellogenin gene expression monitoring programs

USGS Publications Warehouse

Jastrow, Aaron; Gordon, Denise A.; Auger, Kasie M.; Punska, Elizabeth C.; Arcaro, Kathleen F.; Keteles, Kristen; Winkelman, Dana L.; Lattier, David; Biales, Adam; Lazorchak, James M.

2017-01-01

The egg yolk precursor protein vitellogenin is widely used as a biomarker of estrogen exposure in male fish. However, standardized methodology is lacking and little is known regarding the reproducibility of results among laboratories using different equipment, reagents, protocols, and data analysis programs. To address this data gap we tested the reproducibility across laboratories to evaluate vitellogenin gene (vtg) expression and assessed the value of using a freely available software data analysis program. Samples collected from studies of male fathead minnows (Pimephales promelas) exposed to 17α-ethinylestradiol (EE2) and minnows exposed to processed wastewater effluent were evaluated for vtg expression in 4 laboratories. Our results indicate reasonable consistency among laboratories if the free software for expression analysis LinRegPCR is used, with 3 of 4 laboratories detecting vtg in fish exposed to 5 ng/L EE2 (n = 5). All 4 laboratories detected significantly increased vtg levels in 15 male fish exposed to wastewater effluent compared with 15 male fish held in a control stream. Finally, we were able to determine that the source of high interlaboratory variability from complementary deoxyribonucleic acid (cDNA) to quantitative polymerase chain reaction (qPCR) analyses was the expression analysis software unique to each real-time qPCR machine. We successfully eliminated the interlaboratory variability by reanalyzing raw fluorescence data with independent freeware, which yielded cycle thresholds and polymerase chain reaction (PCR) efficiencies that calculated results independently of proprietary software. Our results suggest that laboratories engaged in monitoring programs should validate their PCR protocols and analyze their gene expression data following the guidelines established in the present study for all gene expression biomarkers. 

Interlaboratory trial for the measurement of total cobalt in equine urine and plasma by ICP-MS.

PubMed

Popot, Marie-Agnes; Ho, Emmie N M; Stojiljkovic, Natali; Bagilet, Florian; Remy, Pierre; Maciejewski, Pascal; Loup, Benoit; Chan, George H M; Hargrave, Sabine; Arthur, Rick M; Russo, Charlie; White, James; Hincks, Pamela; Pearce, Clive; Ganio, George; Zahra, Paul; Batty, David; Jarrett, Mark; Brooks, Lydia; Prescott, Lise-Anne; Bailly-Chouriberry, Ludovic; Bonnaire, Yves; Wan, Terence S M

2017-09-01

Cobalt is an essential mineral micronutrient and is regularly present in equine nutritional and feed supplements. Therefore, cobalt is naturally present at low concentrations in biological samples. The administration of cobalt chloride is considered to be blood doping and is thus prohibited. To control the misuse of cobalt, it was mandatory to establish an international threshold for cobalt in plasma and/or in urine. To achieve this goal, an international collaboration, consisting of an interlaboratory comparison between 5 laboratories for the urine study and 8 laboratories for the plasma study, has been undertaken. Quantification of cobalt in the biological samples was performed by inductively coupled plasma-mass spectrometry (ICP-MS). Ring tests were based on the analysis of 5 urine samples supplemented at concentrations ranging from 5 up to 500 ng/mL and 5 plasma samples spiked at concentrations ranging from 0.5 up to 25 ng/mL. The results obtained from the different laboratories were collected, compiled, and compared to assess the reproducibility and robustness of cobalt quantification measurements. The statistical approach for the ring test for total cobalt in urine was based on the determination of percentage deviations from the calculated means, while robust statistics based on the calculated median were applied to the ring test for total cobalt in plasma. The inter-laboratory comparisons in urine and in plasma were successful so that 97.6% of the urine samples and 97.5% of the plasma samples gave satisfactory results. Threshold values for cobalt in plasma and urine were established from data only obtained by laboratories involved in the ring test. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

THE DETERMINATION OF TOTAL ORGANIC HALIDE IN WATER: AN INTERLABORATORY COMPARATIVE STUDY OF TWO METHODS

EPA Science Inventory

Total organic halide (TOX) analyzers are commonly used to measure the amount of dissolved halogenated organic byproducts in disinfected waters. Because of the lack of information on the identity of disinfection byproducts, rigorous testing of the dissolved organic halide (DOX) pr...

An interlaboratory study measuring sex steroids with RIAs and/or ELISAs: Are we comparing apples to oranges?

EPA Science Inventory

Endocrine-disrupting chemicals (EDCs) are exogenous substances released into the environment that can lead to adverse reproductive effects in fish by a number of mechanisms including altering circulating levels of estradiol (E2), testosterone (T) and 11-ketotestosterone (11KT). ...

AN INTERLABORATORY STUDY ON THE USE OF STEROID HORMONES IN EVALUATING ENDOCRINE DISRUPTION

EPA Science Inventory

In recent years, there has been an increased use of the measurement of sex steroid hormone levels in the blood of animals exposed to chemicals as an indicator of reproductive impairment or an alteration in endocrine function. Although levels of hormones are often compared among a...

INTERLABORATORY STUDY ON THE USE OF STEROID HORMONES IN EXAMINING ENDOCRINE DISRUPTION.

EPA Science Inventory

In recent years, there has been an increased use of the measurement of sex steroid hormone levels in the blood of animals exposed to chemicals as an indicator of reproductive impairment or an alteration in endocrine function. Although levels of hormones are often compared among ...

AN INTERLABORATORY STUDY ON THE USE OF STERIOD HORMONES IN EXAMINING ENDOCRINE DISRUPTION.

EPA Science Inventory

In recent years, there has been an increased use of the measurement of sex steroid hormone levels in the blood of animals exposed to chemicals as an indicator of reproductive impairment or an alteration in endocrine function. Although levels of hormones are often compared among a...

How consistent are we? Inter-laboratory comparison for male fathead minnows exposed to 17α-ethinylestradiol

EPA Science Inventory

Transcriptomic approaches are widely used to examine effects of aquatic contaminants in both laboratory and field studies. Fundamental questions remain however for defining the limits of the technology and how it may be used in environmental monitoring programs. Uncertainties exi...

Methods to study legionella transcriptome in vitro and in vivo.

PubMed

Faucher, Sebastien P; Shuman, Howard A

2013-01-01

The study of transcriptome responses can provide insight into the regulatory pathways and genetic factors that contribute to a specific phenotype. For bacterial pathogens, it can identify putative new virulence systems and shed light on the mechanisms underlying the regulation of virulence factors. Microarrays have been previously used to study gene regulation in Legionella pneumophila. In the past few years a sharp reduction of the costs associated with microarray experiments together with the availability of relatively inexpensive custom-designed commercial microarrays has made microarray technology an accessible tool for the majority of researchers. Here we describe the methodologies to conduct microarray experiments from in vitro and in vivo samples.

Interlaboratory comparability, bias, and precision for four laboratories measuring analytes in wet deposition, October 1983-December 1984

USGS Publications Warehouse

Brooks, Myron H.; Schroder, LeRoy J.; Willoughby, Timothy C.

1987-01-01

Four laboratories involved in the routine analysis of wet-deposition samples participated in an interlaboratory comparison program managed by the U.S. Geological Survey. The four participants were: Illinois State Water Survey central analytical laboratory in Champaign, Illinois; U.S. Geological Survey national water-quality laboratories in Atlanta, Georgia, and Denver, Colorado; and Inland Waters Directorate national water-quality laboratory in Burlington, Ontario, Canada. Analyses of interlaboratory samples performed by the four laboratories from October 1983 through December 1984 were compared.Participating laboratories analyzed three types of interlaboratory samples--natural wet deposition, simulated wet deposition, and deionized water--for pH and specific conductance, and for dissolved calcium, magnesium, sodium, sodium, potassium, chloride, sulfate, nitrate, ammonium, and orthophosphate. Natural wet-deposition samples were aliquots of actual wet-deposition samples. Analyses of these samples by the four laboratories were compared using analysis of variance. Test results indicated that pH, calcium, nitrate, and ammonium results were not directly comparable among the four laboratories. Statistically significant differences between laboratory results probably only were meaningful for analyses of dissolved calcium. Simulated wet-deposition samples with known analyte concentrations were used to test each laboratory for analyte bias. Laboratory analyses of calcium, magnesium, sodium, potassium, chloride, sulfate, and nitrate were not significantly different from the known concentrations of these analytes when tested using analysis of variance. Deionized-water samples were used to test each laboratory for reporting of false positive values. The Illinois State Water Survey Laboratory reported the smallest percentage of false positive values for most analytes. Analyte precision was estimated for each laboratory from results of replicate measurements. In general, the Illinois State Water Survey laboratory achieved the greatest precision, whereas the U.S. Geological Survey laboratories achieved the least precision.

Impact of Uniform Methods on Interlaboratory Antibody Titration Variability: Antibody Titration and Uniform Methods.

PubMed

Bachegowda, Lohith S; Cheng, Yan H; Long, Thomas; Shaz, Beth H

2017-01-01

-Substantial variability between different antibody titration methods prompted development and introduction of uniform methods in 2008. -To determine whether uniform methods consistently decrease interlaboratory variation in proficiency testing. -Proficiency testing data for antibody titration between 2009 and 2013 were obtained from the College of American Pathologists. Each laboratory was supplied plasma and red cells to determine anti-A and anti-D antibody titers by their standard method: gel or tube by uniform or other methods at different testing phases (immediate spin and/or room temperature [anti-A], and/or anti-human globulin [AHG: anti-A and anti-D]) with different additives. Interlaboratory variations were compared by analyzing the distribution of titer results by method and phase. -A median of 574 and 1100 responses were reported for anti-A and anti-D antibody titers, respectively, during a 5-year period. The 3 most frequent (median) methods performed for anti-A antibody were uniform tube room temperature (147.5; range, 119-159), uniform tube AHG (143.5; range, 134-150), and other tube AHG (97; range, 82-116); for anti-D antibody, the methods were other tube (451; range, 431-465), uniform tube (404; range, 382-462), and uniform gel (137; range, 121-153). Of the larger reported methods, uniform gel AHG phase for anti-A and anti-D antibodies had the most participants with the same result (mode). For anti-A antibody, 0 of 8 (uniform versus other tube room temperature) and 1 of 8 (uniform versus other tube AHG), and for anti-D antibody, 0 of 8 (uniform versus other tube) and 0 of 8 (uniform versus other gel) proficiency tests showed significant titer variability reduction. -Uniform methods harmonize laboratory techniques but rarely reduce interlaboratory titer variance in comparison with other methods.

Use of allogeneic apheresis stem cell products as an interlaboratory proficiency challenge.

PubMed

Cooling, Laura; Roxbury, Kelly; Hoffmann, Sandra; DeBusscher, Joan; Kota, Usha; Goldstein, Steven; Davenport, Robertson

2017-06-01

AABB Standards requires that laboratories participate in a proficiency test (PT) program for critical analytes. Institutions can purchase commercial PT materials; however, PT can also be performed through interlaboratory exchange. We investigated the utility of allogeneic hematopoietic progenitor cell apheresis (HPC-A) products as an interlaboratory PT challenge for total nucleated cell count (TNC) and CD34 assessment. Three-year retrospective and comparative review of unrelated allogeneic HPC-A products received by the University of Michigan between January 2011 and December 2013. Internal TNC and CD34 count were compared to the external collecting facility by paired t test and linear regression. The absolute and percent difference between external and internal counts and 95% limits of agreeability (95% LA) were determined. Results were analyzed relative to donor center location (international, domestic), time zone (domestic), and calendar year. There was a strong correlation between internal and external TNC, regardless of donor center location or year. For CD34, there was a good correlation between centers (R = 0.88-0.91; slope = 0.95-0.98x) with a median difference of -1% (95% LA, -50%, +47%). This was considerably better than commercial PT challenges, which showed a persistent negative bias for absolute CD34 and CD3 counts. Allogeneic HPC-A products represent an interlaboratory PT exchange for all critical analytes, including TNC and CD34 count, cell viability, and sterility. Allogeneic HPC-A products, which are fresh and transported under validated conditions, are less subject to preanalytical variables that may impact commercial PT samples such as aliquoting and sample homogeneity, commercial additives, and sample stability during manufacturing and transport. © 2017 AABB.

Large-Scale Interlaboratory Study to Develop, Analytically Validate and Apply Highly Multiplexed, Quantitative Peptide Assays to Measure Cancer-Relevant Proteins in Plasma*

PubMed Central

Abbatiello, Susan E.; Schilling, Birgit; Mani, D. R.; Zimmerman, Lisa J.; Hall, Steven C.; MacLean, Brendan; Albertolle, Matthew; Allen, Simon; Burgess, Michael; Cusack, Michael P.; Gosh, Mousumi; Hedrick, Victoria; Held, Jason M.; Inerowicz, H. Dorota; Jackson, Angela; Keshishian, Hasmik; Kinsinger, Christopher R.; Lyssand, John; Makowski, Lee; Mesri, Mehdi; Rodriguez, Henry; Rudnick, Paul; Sadowski, Pawel; Sedransk, Nell; Shaddox, Kent; Skates, Stephen J.; Kuhn, Eric; Smith, Derek; Whiteaker, Jeffery R.; Whitwell, Corbin; Zhang, Shucha; Borchers, Christoph H.; Fisher, Susan J.; Gibson, Bradford W.; Liebler, Daniel C.; MacCoss, Michael J.; Neubert, Thomas A.; Paulovich, Amanda G.; Regnier, Fred E.; Tempst, Paul; Carr, Steven A.

2015-01-01

There is an increasing need in biology and clinical medicine to robustly and reliably measure tens to hundreds of peptides and proteins in clinical and biological samples with high sensitivity, specificity, reproducibility, and repeatability. Previously, we demonstrated that LC-MRM-MS with isotope dilution has suitable performance for quantitative measurements of small numbers of relatively abundant proteins in human plasma and that the resulting assays can be transferred across laboratories while maintaining high reproducibility and quantitative precision. Here, we significantly extend that earlier work, demonstrating that 11 laboratories using 14 LC-MS systems can develop, determine analytical figures of merit, and apply highly multiplexed MRM-MS assays targeting 125 peptides derived from 27 cancer-relevant proteins and seven control proteins to precisely and reproducibly measure the analytes in human plasma. To ensure consistent generation of high quality data, we incorporated a system suitability protocol (SSP) into our experimental design. The SSP enabled real-time monitoring of LC-MRM-MS performance during assay development and implementation, facilitating early detection and correction of chromatographic and instrumental problems. Low to subnanogram/ml sensitivity for proteins in plasma was achieved by one-step immunoaffinity depletion of 14 abundant plasma proteins prior to analysis. Median intra- and interlaboratory reproducibility was <20%, sufficient for most biological studies and candidate protein biomarker verification. Digestion recovery of peptides was assessed and quantitative accuracy improved using heavy-isotope-labeled versions of the proteins as internal standards. Using the highly multiplexed assay, participating laboratories were able to precisely and reproducibly determine the levels of a series of analytes in blinded samples used to simulate an interlaboratory clinical study of patient samples. Our study further establishes that LC-MRM-MS using stable isotope dilution, with appropriate attention to analytical validation and appropriate quality control measures, enables sensitive, specific, reproducible, and quantitative measurements of proteins and peptides in complex biological matrices such as plasma. PMID:25693799

Ultra-trace analysis of 41Ca in urine by accelerator mass spectrometry: an inter-laboratory comparison

PubMed Central

Jackson, George S.; Hillegonds, Darren J.; Muzikar, Paul; Goehring, Brent

2013-01-01

A 41Ca interlaboratory comparison between Lawrence Livermore National Laboratory (LLNL) and the Purdue Rare Isotope Laboratory (PRIME Lab) has been completed. Analysis of the ratios assayed by accelerator mass spectrometry (AMS) shows that there is no statistically significant difference in the ratios. Further, Bayesian analysis shows that the uncertainties reported by both facilities are correct with the possibility of a slight under-estimation by one laboratory. Finally, the chemistry procedures used by the two facilities to produce CaF2 for the cesium sputter ion source are robust and don't yield any significant differences in the final result. PMID:24179312

Results of the first provisional technical secretariat interlaboratory comparison test

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stuff, J.R.; Hoffland, L.

1995-06-01

The principal task of this laboratory in the first Provisional Technical Secretariat (PTS) Interlaboratory Comparison Test was to verify and test the extraction and preparation procedures outlined in the Recommended Operating Procedures for Sampling and Analysis in the Verification of Chemical Disarmament in addition to our laboratory extraction methods and our laboratory analysis methods. Sample preparation began on 16 May 1994 and analysis was completed on 12 June 1994. The analytical methods used included NMR ({sup 1}H and {sup 31}P) GC/AED, GC/MS (EI and methane CI), GC/IRD, HPLC/IC, HPLC/TSP/MS, MS/MS(Electrospray), and CZE.

Toward Improvements in Inter-laboratory Calibration of Argon Isotope Measurements

NASA Astrophysics Data System (ADS)

Hemming, S. R.; Deino, A. L.; Heizler, M. T.; Hodges, K. V.; McIntosh, W. C.; Renne, P. R.; Swisher, C. C., III; Turrin, B. D.; Van Soest, M. C.

2015-12-01

It is important to continue to develop strategies to improve our ability to compare results between laboratories chronometers. The U-Pb community has significantly reduced inter-laboratory biases with the application of a community tracer solution and the distribution of synthetic zircon solutions. Inevitably sample selection and processing and even biases in interpretations will still lead to some disagreements in the assignment of ages. Accordingly natural samples that are shared will be important for achievement of the highest levels of agreement. Analogous improvements in quality and inter-laboratory agreement of analytical aspects of Ar-Ar can be achieved through development of synthetic age standards in gas canisters with multiple pipettes to deliver various controlled amounts of argon to the mass spectrometer. A preliminary proof-of concept comes from the inter-laboratory calibration experiment for the 40Ar/39Ar community. This portable Argon Pipette Intercalibration System (APIS) consists of three 2.7 L canisters each equipped with three pipettes of 0.1, 0.2 and 0.4 cc volumes. The currently traveling APIS has the three canisters filled with air and 40Ar*/39Ar of 1.73 and canister 2 has a 40Ar*/39Ar of 40.98 (~ Alder Creek and Fish Canyon in the same irradiation). With these pipettes it is possible to combine them to provide 0.1, 0.2, 0.3 (0.1+0.2), 0.4, 0.5 (0.1+0.4), 0.6 (0.2+0.4), and 0.7 (0.1+0.2+0.4) cc. The configuration allows a simple test for inter-laboratory biases and for volume/pressure dependent mass fractionation on the measured ratios for a gas with a single argon isotope composition. Although not yet tested, it is also possible to mix gas from any one of the three canisters in proportions of these increments, allowing even more tightly controlled calibration of measurements. We suggest that ultimately each EARTHTIME lab should be equipped with such a system permanently, with a community plan for a traveling system to periodically repeat the inter-calibration tests. The composition(s) of such systems may not be the same for each lab, depending on the requirements of equipment and main age ranges targeted. But with a relatively small number of end member compositions it should be possible to greatly improve the calibration capability of the community.

AACCI Approved Methods Technical Committee Report: Collaborative study on a method for determining the water holding capacity of pulse flours and their protein materials (AACCI Method 56-37.01)

USDA-ARS?s Scientific Manuscript database

A method for determining water holding capacity (WHC) of pulse flours and protein materials has been developed and subjected to an interlaboratory study. Eleven participants analyzed twelve blind duplicates of six different samples in a collaborative study to evaluate the repeatability and reproduci...

Evaluation of platelet adhesion and activation on polymers: Round-robin study to assess inter-center variability.

PubMed

Braune, S; Sperling, C; Maitz, M F; Steinseifer, U; Clauser, J; Hiebl, B; Krajewski, S; Wendel, H P; Jung, F

2017-10-01

The regulatory agencies provide recommendations rather than protocols or standard operation procedures for the hemocompatibility evaluation of novel materials e.g. for cardiovascular applications. Thus, there is a lack of specifications with regard to test setups and procedures. As a consequence, laboratories worldwide perform in vitro assays under substantially different test conditions, so that inter-laboratory and inter-study comparisons are impossible. Here, we report about a prospective, randomized and double-blind multicenter trial which demonstrates that standardization of in vitro test protocols allows a reproducible assessment of platelet adhesion and activation from fresh human platelet rich plasma as possible indicators of the thrombogenicity of cardiovascular implants. Standardization of the reported static in vitro setup resulted in a laboratory independent scoring of the following materials: poly(dimethyl siloxane) (PDMS), poly(ethylene terephthalate) (PET) and poly(tetrafluoro ethylene) (PTFE). The results of this in vitro study provide evidence that inter-laboratory and inter-study comparisons can be achieved for the evaluation of the adhesion and activation of platelets on blood-contacting biomaterials by stringent standardization of test protocols. Copyright © 2017 Elsevier B.V. All rights reserved.

The MicroArray Quality Control (MAQC)-II study of common practices for the development and validation of microarray-based predictive models

EPA Science Inventory

The second phase of the MicroArray Quality Control (MAQC-II) project evaluated common practices for developing and validating microarray-based models aimed at predicting toxicological and clinical endpoints. Thirty-six teams developed classifiers for 13 endpoints - some easy, som...

Rheology measurements of a biomass slurry : an inter-laboratory study

Treesearch

Jonathan J. Stickel; Jeffrey S. Knutsen; Matthew W. Liberatore; Wing Luu; Douglas W. Bousfield; Daniel J. Klingenberg; Tim Scott; Thatcher W. Root; Max R. Ehrhardt; Thomas O. Monz

2009-01-01

The conversion of biomass, specifically lignocellulosic biomass, into fuels and chemicals has recently gained national attention as an alternative to the use of fossil fuels. Increasing the concentration of the biomass solids during biochemical conversion has a large potential to reduce production costs. These concentrated biomass slurries have highly viscous, non-...

More rapid edgewise crush test methods

Treesearch

Thomas J. Urbanik; Arthur H. Catlin; Davide R. Friedman; Richard C. Lund

1993-01-01

The use of paraffin wax to reinforce the loading edges of corrugated fiberboard edge-crush specimens requires that the specimens be reconditioned after waxing. The traditional practice employing a 24-h reconditioning period is a conservative approach based on the moisture response rate of corrugated containers. An interlaboratory study was conducted to determine the...

Interlaboratory testing of 42-d Hyalella azteca survival, growth and reproduction method with sediment and water-only exposures

EPA Science Inventory

Over the past four years, USEPA-Duluth, USGS-Columbia, the Illinois Natural History Survey, and Environment Canada have conducted studies to refine the USEPA/ASTM International methods for conducting 10- to 42-d water or sediment toxicity exposures with Hyalella azteca. In advanc...

POSSIBLE APPROACHES FOR THE ESTABLISHMENT OF INTERLABORATORY COMPARABILITY IN THE DETERMINATION OF POLYCHLORINATED BIPHENYLS IN HUMAN SERUM

EPA Science Inventory

The Massachusetts Department of Public Health, with the assistance of the centers for Disease Control, conducted a study to determine the prevalence of elevated levels (>30ppb) of polychlorinated biphenyls (PCBs) in serum taken from residents of the greater New Bedford area in Ma...

Interlaboratory validation of quantitative duplex real-time PCR method for screening analysis of genetically modified maize.

PubMed

Takabatake, Reona; Koiwa, Tomohiro; Kasahara, Masaki; Takashima, Kaori; Futo, Satoshi; Minegishi, Yasutaka; Akiyama, Hiroshi; Teshima, Reiko; Oguchi, Taichi; Mano, Junichi; Furui, Satoshi; Kitta, Kazumi

2011-01-01

To reduce the cost and time required to routinely perform the genetically modified organism (GMO) test, we developed a duplex quantitative real-time PCR method for a screening analysis simultaneously targeting an event-specific segment for GA21 and Cauliflower Mosaic Virus 35S promoter (P35S) segment [Oguchi et al., J. Food Hyg. Soc. Japan, 50, 117-125 (2009)]. To confirm the validity of the method, an interlaboratory collaborative study was conducted. In the collaborative study, conversion factors (Cfs), which are required to calculate the GMO amount (%), were first determined for two real-time PCR instruments, the ABI PRISM 7900HT and the ABI PRISM 7500. A blind test was then conducted. The limit of quantitation for both GA21 and P35S was estimated to be 0.5% or less. The trueness and precision were evaluated as the bias and reproducibility of the relative standard deviation (RSD(R)). The determined bias and RSD(R) were each less than 25%. We believe the developed method would be useful for the practical screening analysis of GM maize.

Increased frequencies of aberrant sperm as indicators of mutagenic damage in mice.

PubMed

Soares, E R; Sheridan, W; Haseman, J K; Segall, M

1979-02-01

We have tested the effects of TEM in 3 strains of mice using the sperm morphology assay. In addition, we have made an attempt to evaluate this test system with respect to experimental design, statistical problems and possible interlaboratory differences. Treatment with TEM results in significant increases in the percent of abnormally shaped sperm. These increases are readily detectable in sperm treated as spermatocytes and spermatogonial stages. Our data indicate possible problems associated with inter-laboratory variation in slide analysis. We have found that despite the introduction of such sources of variation, our data were consistent with respect to the effects of TEM. Another area of concern in the sperm morphology test is the presence of "outlier" animals. In our study, such animals comprised 4% of the total number of animals considered. Statistical analysis of the slides from these animals have shown that this problem can be dealt with and that when recognized as such, "outliers" do not effect the outcome of the sperm morphology assay.

Transferability and inter-laboratory variability assessment of the in vitro bovine oocyte fertilization test.

PubMed

Tessaro, Irene; Modina, Silvia C; Crotti, Gabriella; Franciosi, Federica; Colleoni, Silvia; Lodde, Valentina; Galli, Cesare; Lazzari, Giovanna; Luciano, Alberto M

2015-01-01

The dramatic increase in the number of animals required for reproductive toxicity testing imposes the validation of alternative methods to reduce the use of laboratory animals. As we previously demonstrated for in vitro maturation test of bovine oocytes, the present study describes the transferability assessment and the inter-laboratory variability of an in vitro test able to identify chemical effects during the process of bovine oocyte fertilization. Eight chemicals with well-known toxic properties (benzo[a]pyrene, busulfan, cadmium chloride, cycloheximide, diethylstilbestrol, ketoconazole, methylacetoacetate, mifepristone/RU-486) were tested in two well-trained laboratories. The statistical analysis demonstrated no differences in the EC50 values for each chemical in within (inter-runs) and in between-laboratory variability of the proposed test. We therefore conclude that the bovine in vitro fertilization test could advance toward the validation process as alternative in vitro method and become part of an integrated testing strategy in order to predict chemical hazards on mammalian fertility. Copyright © 2015 Elsevier Inc. All rights reserved.

GaMin’11 – an international inter-laboratory comparison for geochemical CO₂ - saline fluid - mineral interaction experiments

DOE PAGES

Ostertag-Henning, C.; Risse, A.; Thomas, B.; ...

2014-12-31

Due to the strong interest in geochemical CO₂-fluid-rock interaction in the context of geological storage of CO₂ a growing number of research groups have used a variety of different experimental ways to identify important geochemical dissolution or precipitation reactions and – if possible – quantify the rates and extent of mineral or rock alteration. In this inter-laboratory comparison the gas-fluid-mineral reactions of three samples of rock-forming minerals have been investigated by 11 experimental labs. The reported results point to robust identification of the major processes in the experiments by most groups. The dissolution rates derived from the changes in compositionmore » of the aqueous phase are consistent overall, but the variation could be reduced by using similar corrections for changing parameters in the reaction cells over time. The comparison of experimental setups and procedures as well as of data corrections identified potential improvements for future gas-fluid-rock studies.« less

Trueness and precision of the real-time RT-PCR method for quantifying the chronic bee paralysis virus genome in bee homogenates evaluated by a comparative inter-laboratory study.

PubMed

Schurr, Frank; Cougoule, Nicolas; Rivière, Marie-Pierre; Ribière-Chabert, Magali; Achour, Hamid; Ádám, Dán; Castillo, Carlos; de Graaf, Dirk C; Forsgren, Eva; Granato, Anna; Heinikainen, Sirpa; Jurovčíková, Júlia; Kryger, Per; Manson, Christine; Ménard, Marie-Françoise; Perennes, Stéphane; Schäfer, Marc O; Ibañez, Elena San Miguel; Silva, João; Gajger, Ivana Tlak; Tomkies, Victoria; Toplak, Ivan; Viry, Alain; Zdańska, Dagmara; Dubois, Eric

2017-10-01

The Chronic bee paralysis virus (CBPV) is the aetiological agent of chronic bee paralysis, a contagious disease associated with nervous disorders in adult honeybees leading to massive mortalities in front of the hives. Some of the clinical signs frequently reported, such as trembling, may be confused with intoxication syndromes. Therefore, laboratory diagnosis using real-time PCR to quantify CBPV loads is used to confirm disease. Clinical signs of chronic paralysis are usually associated with viral loads higher than 10 8 copies of CBPV genome copies per bee (8 log 10 CBPV/bee). This threshold is used by the European Union Reference Laboratory for Bee Health to diagnose the disease. In 2015, the accuracy of measurements of three CBPV loads (5, 8 and 9 log 10 CBPV/bee) was assessed through an inter-laboratory study. Twenty-one participants, including 16 European National Reference Laboratories, received 13 homogenates of CBPV-infected bees adjusted to the three loads. Participants were requested to use the method usually employed for routine diagnosis. The quantitative results (n=270) were analysed according to international standards NF ISO 13528 (2015) and NF ISO 5725-2 (1994). The standard deviations of measurement reproducibility (S R ) were 0.83, 1.06 and 1.16 at viral loads 5, 8 and 9 log 10 CBPV/bee, respectively. The inter-laboratory confidence of viral quantification (+/- 1.96S R ) at the diagnostic threshold (8 log 10 CBPV/bee) was+/- 2.08 log 10 CBPV/bee. These results highlight the need to take into account the confidence of measurements in epidemiological studies using results from different laboratories. Considering this confidence, viral loads over 6 log 10 CBPV/bee may be considered to indicate probable cases of chronic paralysis. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Inter-laboratory exercise on antibiotic drugs analysis in aqueous samples.

PubMed

Roig, B; Brogat, M; Mompelat, S; Leveque, J; Cadiere, A; Thomas, O

2012-08-30

An inter-laboratory exercise was organized under the PHARMAS EU project, by the Advanced School of Public Health (EHESP), in order to evaluate the performances of analytical methods for the measurement of antibiotics in waters (surface and tap). This is the first time such an exercise on antibiotics has been organized in Europe, using different kinds of analytical methods and devices. In this exercise thirteen laboratories from five countries (Canada, France, Italy, the Netherlands and Portugal) participated, and a total number of 78 samples were distributed. During the exercise, 2 testing samples (3 bottles of each) prepared from tap water and river water, respectively, spiked with antibiotics, were sent to participants and analyzed over a period of one month. A final number of 77 (98.7%) testing samples were considered. Depending on substances studied by each participant, 305 values in duplicate were collected, with the results for each sample being expressed as the target concentration. A statistical study was initiated using 611 results. The mean value, standard deviation, coefficient of variation, standard uncertainty of the mean, median, the minimum and maximum values of each series as well as the 95% confidence interval were obtained from each participant laboratory. In this exercise, 36 results (6% of accounted values) were outliers according to the distribution over the median (box plot). The outlier results were excluded. In order to establish the stability of testing samples in the course of the exercise, differences between variances obtained for every type of sample at different intervals were evaluated. The results showed no representative variations and it can be considered that all samples were stable during the exercise. The goals of this inter-laboratory study were to assess results variability when analysis is conducted by different laboratories, to evaluate the influence of different matrix samples, and to determine the rate at which participating laboratories successfully completed the tests initiated. Copyright © 2012 Elsevier B.V. All rights reserved.

Genotyping microarray: Mutation screening in Spanish families with autosomal dominant retinitis pigmentosa

PubMed Central

García-Hoyos, María; Cortón, Marta; Ávila-Fernández, Almudena; Riveiro-Álvarez, Rosa; Giménez, Ascensión; Hernan, Inma; Carballo, Miguel; Ayuso, Carmen

2012-01-01

Purpose Presently, 22 genes have been described in association with autosomal dominant retinitis pigmentosa (adRP); however, they explain only 50% of all cases, making genetic diagnosis of this disease difficult and costly. The aim of this study was to evaluate a specific genotyping microarray for its application to the molecular diagnosis of adRP in Spanish patients. Methods We analyzed 139 unrelated Spanish families with adRP. Samples were studied by using a genotyping microarray (adRP). All mutations found were further confirmed with automatic sequencing. Rhodopsin (RHO) sequencing was performed in all negative samples for the genotyping microarray. Results The adRP genotyping microarray detected the mutation associated with the disease in 20 of the 139 families with adRP. As in other populations, RHO was found to be the most frequently mutated gene in these families (7.9% of the microarray genotyped families). The rate of false positives (microarray results not confirmed with sequencing) and false negatives (mutations in RHO detected with sequencing but not with the genotyping microarray) were established, and high levels of analytical sensitivity (95%) and specificity (100%) were found. Diagnostic accuracy was 15.1%. Conclusions The adRP genotyping microarray is a quick, cost-efficient first step in the molecular diagnosis of Spanish patients with adRP. PMID:22736939

Isolation of RNA From Peripheral Blood Cells: A Validation Study for Molecular Diagnostics by Microarray and Kinetic RT-PCR Assays - Application in Aerospace Medicine

DTIC Science & Technology

2004-01-01

of RNA From Peripheral Blood Cells: A Validation Study for Molecular Diagnostics by Microarray and Kinetic RT-PCR Assays  Application in...VALIDATION STUDY FOR MOLECULAR DIAGNOSTICS BY MICROARRAY AND KINETIC RT-PCR ASSAYS  APPLICATION IN AEROSPACE MEDICINE INTRODUCTION Extraction of cellular

Measurement of dissolved organic matter fluorescense in aquatic environments: An interlaboratory comparison

USGS Publications Warehouse

Murphy, Kathleen R.; Butler, Kenna D.; Spencer, Robert G. M.; Stedmon, Colin A.; Boehme, Jennifer R.; Aiken, George R.

2010-01-01

The fluorescent properties of dissolved organic matter (DOM) are often studied in order to infer DOM characteristics in aquatic environments, including source, quantity, composition, and behavior. While a potentially powerful technique, a single widely implemented standard method for correcting and presenting fluorescence measurements is lacking, leading to difficulties when comparing data collected by different research groups. This paper reports on a large-scale interlaboratory comparison in which natural samples and well-characterized fluorophores were analyzed in 20 laboratories in the U.S., Europe, and Australia. Shortcomings were evident in several areas, including data quality-assurance, the accuracy of spectral correction factors used to correct EEMs, and the treatment of optically dense samples. Data corrected by participants according to individual laboratory procedures were more variable than when corrected under a standard protocol. Wavelength dependency in measurement precision and accuracy were observed within and between instruments, even in corrected data. In an effort to reduce future occurrences of similar problems, algorithms for correcting and calibrating EEMs are described in detail, and MATLAB scripts for implementing the study's protocol are provided. Combined with the recent expansion of spectral fluorescence standards, this approach will serve to increase the intercomparability of DOM fluorescence studies.

Advances in cell-free protein array methods.

PubMed

Yu, Xiaobo; Petritis, Brianne; Duan, Hu; Xu, Danke; LaBaer, Joshua

2018-01-01

Cell-free protein microarrays represent a special form of protein microarray which display proteins made fresh at the time of the experiment, avoiding storage and denaturation. They have been used increasingly in basic and translational research over the past decade to study protein-protein interactions, the pathogen-host relationship, post-translational modifications, and antibody biomarkers of different human diseases. Their role in the first blood-based diagnostic test for early stage breast cancer highlights their value in managing human health. Cell-free protein microarrays will continue to evolve to become widespread tools for research and clinical management. Areas covered: We review the advantages and disadvantages of different cell-free protein arrays, with an emphasis on the methods that have been studied in the last five years. We also discuss the applications of each microarray method. Expert commentary: Given the growing roles and impact of cell-free protein microarrays in research and medicine, we discuss: 1) the current technical and practical limitations of cell-free protein microarrays; 2) the biomarker discovery and verification pipeline using protein microarrays; and 3) how cell-free protein microarrays will advance over the next five years, both in their technology and applications.

Comparative tests of bench equipment for fuel control system testing of gas-turbine engine

NASA Astrophysics Data System (ADS)

Shendaleva, E. V.

2018-04-01

The relevance of interlaboratory comparative researches is confirmed by attention of world metrological community to this field of activity. Use of the interlaboratory comparative research methodology not only for single gages collation, but also for bench equipment complexes, such as modeling stands for fuel control system testing of gas-turbine engine, is offered. In this case a comparative measure of different bench equipment will be the control fuel pump. Ensuring traceability of measuring result received at test benches of various air enterprises, development and introduction of national standards to practice of bench tests and, eventually, improvement of quality and safety of a aircraft equipment is result of this approach.

An evaluation of two-channel ChIP-on-chip and DNA methylation microarray normalization strategies

PubMed Central

2012-01-01

Background The combination of chromatin immunoprecipitation with two-channel microarray technology enables genome-wide mapping of binding sites of DNA-interacting proteins (ChIP-on-chip) or sites with methylated CpG di-nucleotides (DNA methylation microarray). These powerful tools are the gateway to understanding gene transcription regulation. Since the goals of such studies, the sample preparation procedures, the microarray content and study design are all different from transcriptomics microarrays, the data pre-processing strategies traditionally applied to transcriptomics microarrays may not be appropriate. Particularly, the main challenge of the normalization of "regulation microarrays" is (i) to make the data of individual microarrays quantitatively comparable and (ii) to keep the signals of the enriched probes, representing DNA sequences from the precipitate, as distinguishable as possible from the signals of the un-enriched probes, representing DNA sequences largely absent from the precipitate. Results We compare several widely used normalization approaches (VSN, LOWESS, quantile, T-quantile, Tukey's biweight scaling, Peng's method) applied to a selection of regulation microarray datasets, ranging from DNA methylation to transcription factor binding and histone modification studies. Through comparison of the data distributions of control probes and gene promoter probes before and after normalization, and assessment of the power to identify known enriched genomic regions after normalization, we demonstrate that there are clear differences in performance between normalization procedures. Conclusion T-quantile normalization applied separately on the channels and Tukey's biweight scaling outperform other methods in terms of the conservation of enriched and un-enriched signal separation, as well as in identification of genomic regions known to be enriched. T-quantile normalization is preferable as it additionally improves comparability between microarrays. In contrast, popular normalization approaches like quantile, LOWESS, Peng's method and VSN normalization alter the data distributions of regulation microarrays to such an extent that using these approaches will impact the reliability of the downstream analysis substantially. PMID:22276688

Saccadic Reaction Times in Infants and Adults: Spatiotemporal Factors, Gender, and Interlaboratory Variation

ERIC Educational Resources Information Center

Kenward, Ben; Koch, Felix-Sebastian; Forssman, Ida; Brehm, Julia; Tidemann, Linda; Sundqvist, Annette; Marciszkom, Carin; Hermansen, Tone Kristine; Heimann, Mikael; Gredebäck, Gustaf

2017-01-01

Saccade latency is widely used across infant psychology to investigate infants' understanding of events. Interpreting particular latency values requires knowledge of standard saccadic RTs, but there is no consensus as to typical values. This study provides standard estimates of infants' (n = 194, ages 9 to 15 months) saccadic RTs under a range of…

An inter-laboratory study on the variability in measured concentrations of 17Beta-estradiol, testosterone and 11-ketotestosterone in white sucker: implications and recommendations

EPA Science Inventory

Endocrine-disrupting chemicals (EDCs) are exogenous substances that can lead to impacts on the reproduction of fish sometimes by altering circulating concentrations of 17â-estradiol (E2), testosterone (T) and 11-ketotestosterone (11-KT). Common methods to measure steroids in pla...

A Human Lectin Microarray for Sperm Surface Glycosylation Analysis *

PubMed Central

Sun, Yangyang; Cheng, Li; Gu, Yihua; Xin, Aijie; Wu, Bin; Zhou, Shumin; Guo, Shujuan; Liu, Yin; Diao, Hua; Shi, Huijuan; Wang, Guangyu; Tao, Sheng-ce

2016-01-01

Glycosylation is one of the most abundant and functionally important protein post-translational modifications. As such, technology for efficient glycosylation analysis is in high demand. Lectin microarrays are a powerful tool for such investigations and have been successfully applied for a variety of glycobiological studies. However, most of the current lectin microarrays are primarily constructed from plant lectins, which are not well suited for studies of human glycosylation because of the extreme complexity of human glycans. Herein, we constructed a human lectin microarray with 60 human lectin and lectin-like proteins. All of the lectins and lectin-like proteins were purified from yeast, and most showed binding to human glycans. To demonstrate the applicability of the human lectin microarray, human sperm were probed on the microarray and strong bindings were observed for several lectins, including galectin-1, 7, 8, GalNAc-T6, and ERGIC-53 (LMAN1). These bindings were validated by flow cytometry and fluorescence immunostaining. Further, mass spectrometry analysis showed that galectin-1 binds several membrane-associated proteins including heat shock protein 90. Finally, functional assays showed that binding of galectin-8 could significantly enhance the acrosome reaction within human sperms. To our knowledge, this is the first construction of a human lectin microarray, and we anticipate it will find wide use for a range of human or mammalian studies, alone or in combination with plant lectin microarrays. PMID:27364157

Graph Based Study of Allergen Cross-Reactivity of Plant Lipid Transfer Proteins (LTPs) Using Microarray in a Multicenter Study

PubMed Central

Palacín, Arantxa; Gómez-Casado, Cristina; Rivas, Luis A.; Aguirre, Jacobo; Tordesillas, Leticia; Bartra, Joan; Blanco, Carlos; Carrillo, Teresa; Cuesta-Herranz, Javier; de Frutos, Consolación; Álvarez-Eire, Genoveva García; Fernández, Francisco J.; Gamboa, Pedro; Muñoz, Rosa; Sánchez-Monge, Rosa; Sirvent, Sofía; Torres, María J.; Varela-Losada, Susana; Rodríguez, Rosalía; Parro, Victor; Blanca, Miguel; Salcedo, Gabriel; Díaz-Perales, Araceli

2012-01-01

The study of cross-reactivity in allergy is key to both understanding. the allergic response of many patients and providing them with a rational treatment In the present study, protein microarrays and a co-sensitization graph approach were used in conjunction with an allergen microarray immunoassay. This enabled us to include a wide number of proteins and a large number of patients, and to study sensitization profiles among members of the LTP family. Fourteen LTPs from the most frequent plant food-induced allergies in the geographical area studied were printed into a microarray specifically designed for this research. 212 patients with fruit allergy and 117 food-tolerant pollen allergic subjects were recruited from seven regions of Spain with different pollen profiles, and their sera were tested with allergen microarray. This approach has proven itself to be a good tool to study cross-reactivity between members of LTP family, and could become a useful strategy to analyze other families of allergens. PMID:23272072

PRACTICAL STRATEGIES FOR PROCESSING AND ANALYZING SPOTTED OLIGONUCLEOTIDE MICROARRAY DATA

EPA Science Inventory

Thoughtful data analysis is as important as experimental design, biological sample quality, and appropriate experimental procedures for making microarrays a useful supplement to traditional toxicology. In the present study, spotted oligonucleotide microarrays were used to profile...

Development and inter-laboratory assessment of droplet digital PCR assays for multiplex quantification of 15 genetically modified soybean lines.

PubMed

Košir, Alexandra Bogožalec; Spilsberg, Bjørn; Holst-Jensen, Arne; Žel, Jana; Dobnik, David

2017-08-17

Quantification of genetically modified organisms (GMOs) in food and feed products is often required for their labelling or for tolerance thresholds. Standard-curve-based simplex quantitative polymerase chain reaction (qPCR) is the prevailing technology, which is often combined with screening analysis. With the rapidly growing number of GMOs on the world market, qPCR analysis becomes laborious and expensive. Innovative cost-effective approaches are therefore urgently needed. Here, we report the development and inter-laboratory assessment of multiplex assays to quantify GMO soybean using droplet digital PCR (ddPCR). The assays were developed to facilitate testing of foods and feed for compliance with current GMO regulations in the European Union (EU). Within the EU, the threshold for labelling is 0.9% for authorised GMOs per ingredient. Furthermore, the EU has set a technical zero tolerance limit of 0.1% for certain unauthorised GMOs. The novel multiplex ddPCR assays developed target 11 GMO soybean lines that are currently authorised, and four that are tolerated, pending authorisation in the EU. Potential significant improvements in cost efficiency are demonstrated. Performance was assessed for the critical parameters, including limits of detection and quantification, and trueness, repeatability, and robustness. Inter-laboratory performance was also determined on a number of proficiency programme and real-life samples.

Establishment and validation of analytical reference panels for the standardization of quantitative BCR-ABL1 measurements on the international scale.

PubMed

White, Helen E; Hedges, John; Bendit, Israel; Branford, Susan; Colomer, Dolors; Hochhaus, Andreas; Hughes, Timothy; Kamel-Reid, Suzanne; Kim, Dong-Wook; Modur, Vijay; Müller, Martin C; Pagnano, Katia B; Pane, Fabrizio; Radich, Jerry; Cross, Nicholas C P; Labourier, Emmanuel

2013-06-01

Current guidelines for managing Philadelphia-positive chronic myeloid leukemia include monitoring the expression of the BCR-ABL1 (breakpoint cluster region/c-abl oncogene 1, non-receptor tyrosine kinase) fusion gene by quantitative reverse-transcription PCR (RT-qPCR). Our goal was to establish and validate reference panels to mitigate the interlaboratory imprecision of quantitative BCR-ABL1 measurements and to facilitate global standardization on the international scale (IS). Four-level secondary reference panels were manufactured under controlled and validated processes with synthetic Armored RNA Quant molecules (Asuragen) calibrated to reference standards from the WHO and the NIST. Performance was evaluated in IS reference laboratories and with non-IS-standardized RT-qPCR methods. For most methods, percent ratios for BCR-ABL1 e13a2 and e14a2 relative to ABL1 or BCR were robust at 4 different levels and linear over 3 logarithms, from 10% to 0.01% on the IS. The intraassay and interassay imprecision was <2-fold overall. Performance was stable across 3 consecutive lots, in multiple laboratories, and over a period of 18 months to date. International field trials demonstrated the commutability of the reagents and their accurate alignment to the IS within the intra- and interlaboratory imprecision of IS-standardized methods. The synthetic calibrator panels are robust, reproducibly manufactured, analytically calibrated to the WHO primary standards, and compatible with most BCR-ABL1 RT-qPCR assay designs. The broad availability of secondary reference reagents will further facilitate interlaboratory comparative studies and independent quality assessment programs, which are of paramount importance for worldwide standardization of BCR-ABL1 monitoring results and the optimization of current and new therapeutic approaches for chronic myeloid leukemia. © 2013 American Association for Clinical Chemistry.

Interlaboratory Evaluation of a Multiplexed High Information Content In Vitro Genotoxicity Assay

PubMed Central

Bryce, Steven M.; Bernacki, Derek T.; Bemis, Jeffrey C.; Spellman, Richard A.; Engel, Maria E.; Schuler, Maik; Lorge, Elisabeth; Heikkinen, Pekka T.; Hemmann, Ulrike; Thybaud, Véronique; Wilde, Sabrina; Queisser, Nina; Sutter, Andreas; Zeller, Andreas; Guérard, Melanie; Kirkland, David; Dertinger, Stephen D.

2017-01-01

We previously described a multiplexed in vitro genotoxicity assay based on flow cytometric analysis of detergent-liberated nuclei that are simultaneously stained with propidium iodide and labeled with fluorescent antibodies against p53, γH2AX, and phospho-histone H3. Inclusion of a known number of microspheres provides absolute nuclei counts. The work described herein was undertaken to evaluate the interlaboratory transferability of this assay, commercially known as MultiFlow™ DNA Damage Kit— p53, γH2AX, Phospho-histone H3. For these experiments seven laboratories studied reference chemicals from a group of 84 representing clastogens, aneugens, and non-genotoxicants. TK6 cells were exposed to chemicals in 96-well plates over a range of concentrations for 24 hrs. At 4 and 24 hrs cell aliquots were added to the MultiFlow reagent mix and following a brief incubation period flow cytometric analysis occurred, in most cases directly from a 96-well plate via a robotic walk-away data acquisition system. Multiplexed response data were evaluated using two analysis approaches, one based on global evaluation factors (i.e., cutoff values derived from all inter-laboratory data), and a second based on multinomial logistic regression that considers multiple biomarkers simultaneously. Both data analysis strategies were devised to categorize chemicals as predominately exhibiting a clastogenic, aneugenic, or non-genotoxic mode of action (MoA). Based on the aggregate 231 experiments that were performed, assay sensitivity, specificity, and concordance in relation to a priori MoA grouping were ≥ 92%. These results are encouraging as they suggest that two distinct data analysis strategies can rapidly and reliably predict new chemicals’ predominant genotoxic MoA based on data from an efficient and transferable multiplexed in vitro assay. PMID:28370322

DNA Microarray for Rapid Detection and Identification of Food and Water Borne Bacteria: From Dry to Wet Lab.

PubMed

Ranjbar, Reza; Behzadi, Payam; Najafi, Ali; Roudi, Raheleh

2017-01-01

A rapid, accurate, flexible and reliable diagnostic method may significantly decrease the costs of diagnosis and treatment. Designing an appropriate microarray chip reduces noises and probable biases in the final result. The aim of this study was to design and construct a DNA Microarray Chip for a rapid detection and identification of 10 important bacterial agents. In the present survey, 10 unique genomic regions relating to 10 pathogenic bacterial agents including Escherichia coli (E.coli), Shigella boydii, Sh.dysenteriae, Sh.flexneri, Sh.sonnei, Salmonella typhi, S.typhimurium, Brucella sp., Legionella pneumophila, and Vibrio cholera were selected for designing specific long oligo microarray probes. For this reason, the in-silico operations including utilization of the NCBI RefSeq database, Servers of PanSeq and Gview, AlleleID 7.7 and Oligo Analyzer 3.1 was done. On the other hand, the in-vitro part of the study comprised stages of robotic microarray chip probe spotting, bacterial DNAs extraction and DNA labeling, hybridization and microarray chip scanning. In wet lab section, different tools and apparatus such as Nexterion® Slide E, Qarray mini spotter, NimbleGen kit, TrayMix TM S4, and Innoscan 710 were used. A DNA microarray chip including 10 long oligo microarray probes was designed and constructed for detection and identification of 10 pathogenic bacteria. The DNA microarray chip was capable to identify all 10 bacterial agents tested simultaneously. The presence of a professional bioinformatician as a probe designer is needed to design appropriate multifunctional microarray probes to increase the accuracy of the outcomes.

Comparison of gene expression microarray data with count-based RNA measurements informs microarray interpretation.

PubMed

Richard, Arianne C; Lyons, Paul A; Peters, James E; Biasci, Daniele; Flint, Shaun M; Lee, James C; McKinney, Eoin F; Siegel, Richard M; Smith, Kenneth G C

2014-08-04

Although numerous investigations have compared gene expression microarray platforms, preprocessing methods and batch correction algorithms using constructed spike-in or dilution datasets, there remains a paucity of studies examining the properties of microarray data using diverse biological samples. Most microarray experiments seek to identify subtle differences between samples with variable background noise, a scenario poorly represented by constructed datasets. Thus, microarray users lack important information regarding the complexities introduced in real-world experimental settings. The recent development of a multiplexed, digital technology for nucleic acid measurement enables counting of individual RNA molecules without amplification and, for the first time, permits such a study. Using a set of human leukocyte subset RNA samples, we compared previously acquired microarray expression values with RNA molecule counts determined by the nCounter Analysis System (NanoString Technologies) in selected genes. We found that gene measurements across samples correlated well between the two platforms, particularly for high-variance genes, while genes deemed unexpressed by the nCounter generally had both low expression and low variance on the microarray. Confirming previous findings from spike-in and dilution datasets, this "gold-standard" comparison demonstrated signal compression that varied dramatically by expression level and, to a lesser extent, by dataset. Most importantly, examination of three different cell types revealed that noise levels differed across tissues. Microarray measurements generally correlate with relative RNA molecule counts within optimal ranges but suffer from expression-dependent accuracy bias and precision that varies across datasets. We urge microarray users to consider expression-level effects in signal interpretation and to evaluate noise properties in each dataset independently.

An Update to the EARTHTIME Argon Intercalibration Pipette System (APIS): Smoking from the Same Pipe

NASA Astrophysics Data System (ADS)

Turrin, B. D.; Swisher, C. C., III; Hemming, S. R.; Renne, P. R.; Deino, A. L.; Hodges, K. V.; Van Soest, M. C.; Heizler, M. T.

2015-12-01

Background: An initial 40Ar/39Ar inter-calibration experiment, using two of the most commonly used 40Ar/39Ar mineral standards, Fish Canyon (FC, ~28.2 Ma) and Alder Creek (AC, ~1.2 Ma) sanidines, revealed significant inter-laboratory inconsistencies. The reported ages for the AC sanidines range from 1.173 to 1.200 Ma (FC 28.02) (±~2%), ~4 times greater than the reported precisions. These experiments have motivated the 40Ar/39Ar community to reevaluate procedures and subsequent informal lab intercalibrations experiments are in better agreement, but discrepancies remain that need to be addressed. Experiment: In an effort to isolate the causes of these inconsistencies, an Argon Intercalibration Pipette System (APIS) was designed and constructed. The APIS system consists of three gas canisters; one containing atmospheric Ar and the other two canisters contain 40Ar/39Ar ratios that represent FC and AC. The volumes of the pipettes, bulbs and manifold are determined to within 0.4% and both systems were initially filled to the same pressure with Ar standard gases. Each canister has 4x10-10 moles of 40Ar, is equipped with a 0.1, 0.2 and 0.4 cc pipettes and can deliver increments from 0.1-0.7 cc. APIS-1 was designated as the traveling unit that is brought to participating labs, APIS-2 is the reserve/master standard. Early Results and Impressions: APIS-1 has been to four labs (Rutgers, LDEO, New Mexico Tech, and BGC) and is heading to ASU. Early APIS experimental data indicate that the inter-laboratory 40Ar/39Ar age results can meet or exceed the EARTHTIME goal of ±1‰ precision. The inter-laboratory comparisons are ongoing, and will include additional laboratories of opportunity. Lastly, the development of additional mineral standards that "fill in" the age gaps between the existing mineral standards would significantly improve attempting to achieve interlaboratory agreement at the ±1‰ level

Argon Intercalibration Pipette System (APIS): Smoking from the Same Pipe

NASA Astrophysics Data System (ADS)

Turrin, B. D.; Swisher, C. C., III; Hemming, S. R.; Renne, P. R.; Deino, A. L.; Hodges, K. V.; Van Soest, M. C.; Heizler, M. T.

2014-12-01

40Ar/39Ar age inter-calibration experiments, conducted as part of the US NSF sponsored EARTHTIME initiative, (http://www.earth-time.org), using two of the most commonly used 40Ar/39Ar mineral standards, Fish Canyon (FC, ~28.2 Ma) and Alder Creek (AC, ~1.2 Ma) sanidines, have revealed significant inter-laboratory inconsistencies. The reported ages for the AC sanidines range from 1.173 to 1.200 Ma (FC 28.02) (±~2%), ~4 times greater than the reported precision. These experiments have caused the 40Ar/39Ar community to scrutinize procedures and several informal lab intercalibrations have been conducted among different labs. This exercise is leading to better agreement, but discrepancies remain that need to be addressed. In an effort to isolate the cause(s) of these inconsistencies, two Argon Inter-calibration Pipette System (APIS) were designed and constructed. Each consists of three gas canisters; one contains atmospheric Ar, while the other two contain artificial gas mixtures with 40Ar/39Ar ratios similar to those of FC and AC. Each canister has 4x10-10 moles of 40Ar, is equipped with 0.1, 0.2 and 0.4 cc pipettes, and can deliver gas volumes from 0.1-0.7 cc. All volumes were determined manometrically to 0.4% or better and then filled to uniform pressure with Ar standard gases. This experimental design eliminates sample heterogeneity, leaving only interlaboratory variations in gas purification, data reduction, and isotopic measurement as potential sources of interlaboratory calibration discrepancies. APIS-1 was designated as a traveling unit that is brought to participating labs. APIS-2 is the reserve/master standard. Currently, APIS-1 is in its early stages in the voyage and has been to three labs (Rutgers, LDEO, and New Mexico Tech) as of this writing. The interlaboratory comparisons are ongoing, and will include ASU, BGC, Univ. of Wisconsin, and Oregon State University, plus additional laboratories of opportunity. A progress report will be presented at AGU.

Clustering-based spot segmentation of cDNA microarray images.

PubMed

Uslan, Volkan; Bucak, Ihsan Ömür

2010-01-01

Microarrays are utilized as that they provide useful information about thousands of gene expressions simultaneously. In this study segmentation step of microarray image processing has been implemented. Clustering-based methods, fuzzy c-means and k-means, have been applied for the segmentation step that separates the spots from the background. The experiments show that fuzzy c-means have segmented spots of the microarray image more accurately than the k-means.

Inter-laboratory consistency and variability in the buccal micronucleus cytome assay depends on biomarker scored and laboratory experience: results from the HUMNxl international inter-laboratory scoring exercise.

PubMed

Bolognesi, Claudia; Knasmueller, Siegfried; Nersesyan, Armen; Roggieri, Paola; Ceppi, Marcello; Bruzzone, Marco; Blaszczyk, Ewa; Mielzynska-Svach, Danuta; Milic, Mirta; Bonassi, Stefano; Benedetti, Danieli; Da Silva, Juliana; Toledo, Raphael; Salvadori, Daisy Maria Fávero; Groot de Restrepo, Helena; Filipic, Metka; Hercog, Klara; Aktas, Ayça; Burgaz, Sema; Kundi, Michael; Grummt, Tamara; Thomas, Philip; Hor, Maryam; Escudero-Fung, Maria; Holland, Nina; Fenech, Michael

2017-03-01

The buccal micronucleus cytome (BMNcyt) assay in uncultured exfoliated epithelial cells from oral mucosa is widely applied in biomonitoring human exposures to genotoxic agents and is also proposed as a suitable test for prescreening and follow-up of precancerous oral lesions. The main limitation of the assay is the large variability observed in the baseline values of micronuclei (MNi) and other nuclear anomalies mainly related to different scoring criteria. The aim of this international collaborative study, involving laboratories with different level of experience, was to evaluate the inter- and intra-laboratory variations in the BMNcyt parameters, using recently implemented guidelines, in scoring cells from the same pooled samples obtained from healthy subjects (control group) and from cancer patients undergoing radiotherapy (treated group). The results indicate that all laboratories correctly discriminated samples from the two groups by a significant increase of micronucleus (MN) and nuclear bud (NBUD) frequencies and differentiated binucleated (BN) cells, associated with the exposure to ionizing radiation. The experience of the laboratories was shown to play an important role in the identification of the different cell types and nuclear anomalies. MN frequency in differentiated mononucleated (MONO) and BN cells showed the greatest consistency among the laboratories and low variability was also detected in the frequencies of MONO and BN cells. A larger variability was observed in classifying the different cell types, indicating the subjectivity in the interpretation of some of the scoring criteria while reproducibility of the results between scoring sessions was very good. An inter-laboratory calibration exercise is strongly recommended before starting studies with BMNcyt assay involving multiple research centers. © The Author 2016. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Validation of a quantitative cerebrospinal fluid alpha-synuclein assay in a European-wide interlaboratory study.

PubMed

Kruse, Niels; Persson, Staffan; Alcolea, Daniel; Bahl, Justyna M C; Baldeiras, Ines; Capello, Elisabetta; Chiasserini, Davide; Bocchio Chiavetto, Luisella; Emersic, Andreja; Engelborghs, Sebastiaan; Eren, Erden; Fladby, Tormod; Frisoni, Giovanni; García-Ayllón, María-Salud; Genc, Sermin; Gkatzima, Olymbia; Heegaard, Niels H H; Janeiro, André M; Kováčech, Branislav; Kuiperij, H Bea; Leitão, Maria J; Lleó, Alberto; Martins, Madalena; Matos, Mafalda; Mollergard, Hanne M; Nobili, Flavio; Öhrfelt, Annika; Parnetti, Lucilla; de Oliveira, Catarina Resende; Rot, Uros; Sáez-Valero, Javier; Struyfs, Hanne; Tanassi, Julia T; Taylor, Peggy; Tsolaki, Magda; Vanmechelen, Eugeen; Verbeek, Marcel M; Zilka, Norbert; Blennow, Kaj; Zetterberg, Henrik; Mollenhauer, Brit

2015-09-01

Decreased levels of alpha-synuclein (aSyn) in cerebrospinal fluid (CSF) in Parkinson's disease and related synucleinopathies have been reported, however, not consistently in all cross-sectional studies. To test the performance of one recently released human-specific enzyme-linked immunosorbent assay (ELISA) for the quantification of aSyn in CSF, we carried out a round robin trial with 18 participating laboratories trained in CSF ELISA analyses within the BIOMARKAPD project in the EU Joint Program - Neurodegenerative Disease Research. CSF samples (homogeneous aliquots from pools) and ELISA kits (one lot) were provided centrally and data reported back to one laboratory for data analysis. Our study showed that although factors such as preanalytical sample handling and lot-to-lot variability were minimized by our study design, we identified high variation in absolute values of CSF aSyn even when the same samples and same lots of assays were applied. We further demonstrate that although absolute concentrations differ between laboratories the quantitative results are comparable. With further standardization this assay may become an attractive tool for comparing aSyn measurements in diverse settings. Recommendations for further validation experiments and improvement of the interlaboratory results obtained are given. Copyright © 2015 Elsevier Inc. All rights reserved.

The Importance of Normalization on Large and Heterogeneous Microarray Datasets

EPA Science Inventory

DNA microarray technology is a powerful functional genomics tool increasingly used for investigating global gene expression in environmental studies. Microarrays can also be used in identifying biological networks, as they give insight on the complex gene-to-gene interactions, ne...

An Undergraduate Laboratory Exercise to Study the Effect of Darkness on Plant Gene Expression Using DNA Microarray

ERIC Educational Resources Information Center

Chang, Ming-Mei; Briggs, George M.

2007-01-01

DNA microarrays are microscopic arrays on a solid surface, typically a glass slide, on which DNA oligonucleotides are deposited or synthesized in a high-density matrix with a predetermined spatial order. Several types of DNA microarrays have been developed and used for various biological studies. Here, we developed an undergraduate laboratory…

Interlaboratory validation of an improved U.S. Food and Drug Administration method for detection of Cyclospora cayetanensis in produce using TaqMan real-time PCR

USDA-ARS?s Scientific Manuscript database

A collaborative validation study was performed to evaluate the performance of a new U.S. Food and Drug Administration method developed for detection of the protozoan parasite, Cyclospora cayetanensis, on cilantro and raspberries. The method includes a sample preparation step in which oocysts are re...

Development and Interlaboratory Validation of a Simple Screening Method for Genetically Modified Maize Using a ΔΔC(q)-Based Multiplex Real-Time PCR Assay.

PubMed

Noguchi, Akio; Nakamura, Kosuke; Sakata, Kozue; Sato-Fukuda, Nozomi; Ishigaki, Takumi; Mano, Junichi; Takabatake, Reona; Kitta, Kazumi; Teshima, Reiko; Kondo, Kazunari; Nishimaki-Mogami, Tomoko

2016-04-19

A number of genetically modified (GM) maize events have been developed and approved worldwide for commercial cultivation. A screening method is needed to monitor GM maize approved for commercialization in countries that mandate the labeling of foods containing a specified threshold level of GM crops. In Japan, a screening method has been implemented to monitor approved GM maize since 2001. However, the screening method currently used in Japan is time-consuming and requires generation of a calibration curve and experimental conversion factor (C(f)) value. We developed a simple screening method that avoids the need for a calibration curve and C(f) value. In this method, ΔC(q) values between the target sequences and the endogenous gene are calculated using multiplex real-time PCR, and the ΔΔC(q) value between the analytical and control samples is used as the criterion for determining analytical samples in which the GM organism content is below the threshold level for labeling of GM crops. An interlaboratory study indicated that the method is applicable independently with at least two models of PCR instruments used in this study.

The Development of a Novel, Validated, Rapid and Simple Method for the Detection of Sarcocystis fayeri in Horse Meat in the Sanitary Control Setting.

PubMed

Furukawa, Masato; Minegishi, Yasutaka; Izumiyama, Shinji; Yagita, Kenji; Mori, Hideto; Uemura, Taku; Etoh, Yoshiki; Maeda, Eriko; Sasaki, Mari; Ichinose, Kazuya; Harada, Seiya; Kamata, Yoichi; Otagiri, Masaki; Sugita-Konishi, Yoshiko; Ohnishi, Takahiro

2016-01-01

Sarcocystis fayeri (S. fayeri) is a newly identified causative agent of foodborne disease that is associated with the consumption of raw horse meat. The testing methods prescribed by the Ministry of Health, Labour and Welfare of Japan are time consuming and require the use of expensive equipment and a high level of technical expertise. Accordingly, these methods are not suitable for use in the routine sanitary control setting to prevent outbreaks of foodborne disease. In order to solve these problems, we have developed a new, rapid and simple testing method using LAMP, which takes only 1 hour to perform and which does not involve the use of any expensive equipment or expert techniques. For the validation of this method, an inter-laboratory study was performed among 5 institutes using 10 samples infected with various concentrations of S. fayeri. The results of the inter-laboratory study demonstrated that our LAMP method could detect S. fayeri at concentrations greater than 10(4) copies/g. Thus, this new method could be useful in screening for S. fayeri as a routine sanitary control procedure.

Tendency for interlaboratory precision in the GMO analysis method based on real-time PCR.

PubMed

Kodama, Takashi; Kurosawa, Yasunori; Kitta, Kazumi; Naito, Shigehiro

2010-01-01

The Horwitz curve estimates interlaboratory precision as a function only of concentration, and is frequently used as a method performance criterion in food analysis with chemical methods. The quantitative biochemical methods based on real-time PCR require an analogous criterion to progressively promote method validation. We analyzed the tendency of precision using a simplex real-time PCR technique in 53 collaborative studies of seven genetically modified (GM) crops. Reproducibility standard deviation (SR) and repeatability standard deviation (Sr) of the genetically modified organism (GMO) amount (%) was more or less independent of GM crops (i.e., maize, soybean, cotton, oilseed rape, potato, sugar beet, and rice) and evaluation procedure steps. Some studies evaluated whole steps consisting of DNA extraction and PCR quantitation, whereas others focused only on the PCR quantitation step by using DNA extraction solutions. Therefore, SR and Sr for GMO amount (%) are functions only of concentration similar to the Horwitz curve. We proposed S(R) = 0.1971C 0.8685 and S(r) = 0.1478C 0.8424, where C is the GMO amount (%). We also proposed a method performance index in GMO quantitative methods that is analogous to the Horwitz Ratio.

Assessing differential expression in two-color microarrays: a resampling-based empirical Bayes approach.

PubMed

Li, Dongmei; Le Pape, Marc A; Parikh, Nisha I; Chen, Will X; Dye, Timothy D

2013-01-01

Microarrays are widely used for examining differential gene expression, identifying single nucleotide polymorphisms, and detecting methylation loci. Multiple testing methods in microarray data analysis aim at controlling both Type I and Type II error rates; however, real microarray data do not always fit their distribution assumptions. Smyth's ubiquitous parametric method, for example, inadequately accommodates violations of normality assumptions, resulting in inflated Type I error rates. The Significance Analysis of Microarrays, another widely used microarray data analysis method, is based on a permutation test and is robust to non-normally distributed data; however, the Significance Analysis of Microarrays method fold change criteria are problematic, and can critically alter the conclusion of a study, as a result of compositional changes of the control data set in the analysis. We propose a novel approach, combining resampling with empirical Bayes methods: the Resampling-based empirical Bayes Methods. This approach not only reduces false discovery rates for non-normally distributed microarray data, but it is also impervious to fold change threshold since no control data set selection is needed. Through simulation studies, sensitivities, specificities, total rejections, and false discovery rates are compared across the Smyth's parametric method, the Significance Analysis of Microarrays, and the Resampling-based empirical Bayes Methods. Differences in false discovery rates controls between each approach are illustrated through a preterm delivery methylation study. The results show that the Resampling-based empirical Bayes Methods offer significantly higher specificity and lower false discovery rates compared to Smyth's parametric method when data are not normally distributed. The Resampling-based empirical Bayes Methods also offers higher statistical power than the Significance Analysis of Microarrays method when the proportion of significantly differentially expressed genes is large for both normally and non-normally distributed data. Finally, the Resampling-based empirical Bayes Methods are generalizable to next generation sequencing RNA-seq data analysis.

An examination of the hexokinase method for serum glucose assay using external quality assessment data.

PubMed

Westwood, A; Bullock, D G; Whitehead, T P

1986-01-01

Hexokinase methods for serum glucose assay appeared to give slightly but consistently higher inter-laboratory coefficients of variation than all methods combined in the UK External Quality Assessment Scheme; their performance over a two-year period was therefore compared with that for three groups of glucose oxidase methods. This assessment showed no intrinsic inferiority in the hexokinase method. The greater variation may be due to the more heterogeneous group of instruments, particularly discrete analysers, on which the method is used. The Beckman Glucose Analyzer and Astra group (using a glucose oxidase method) showed the least inter-laboratory variability but also the lowest mean value. No comment is offered on the absolute accuracy of any of the methods.

LipidQC: Method Validation Tool for Visual Comparison to SRM 1950 Using NIST Interlaboratory Comparison Exercise Lipid Consensus Mean Estimate Values.

PubMed

Ulmer, Candice Z; Ragland, Jared M; Koelmel, Jeremy P; Heckert, Alan; Jones, Christina M; Garrett, Timothy J; Yost, Richard A; Bowden, John A

2017-12-19

As advances in analytical separation techniques, mass spectrometry instrumentation, and data processing platforms continue to spur growth in the lipidomics field, more structurally unique lipid species are detected and annotated. The lipidomics community is in need of benchmark reference values to assess the validity of various lipidomics workflows in providing accurate quantitative measurements across the diverse lipidome. LipidQC addresses the harmonization challenge in lipid quantitation by providing a semiautomated process, independent of analytical platform, for visual comparison of experimental results of National Institute of Standards and Technology Standard Reference Material (SRM) 1950, "Metabolites in Frozen Human Plasma", against benchmark consensus mean concentrations derived from the NIST Lipidomics Interlaboratory Comparison Exercise.

An interlaboratory study of TEX86 and BIT analysis of sediments, extracts, and standard mixtures

NASA Astrophysics Data System (ADS)

Schouten, Stefan; Hopmans, Ellen C.; Rosell-Melé, Antoni; Pearson, Ann; Adam, Pierre; Bauersachs, Thorsten; Bard, Edouard; Bernasconi, Stefano M.; Bianchi, Thomas S.; Brocks, Jochen J.; Carlson, Laura Truxal; Castañeda, Isla S.; Derenne, Sylvie; Selver, Ayça. Doǧrul; Dutta, Koushik; Eglinton, Timothy; Fosse, Celine; Galy, Valier; Grice, Kliti; Hinrichs, Kai-Uwe; Huang, Yongsong; Huguet, Arnaud; Huguet, Carme; Hurley, Sarah; Ingalls, Anitra; Jia, Guodong; Keely, Brendan; Knappy, Chris; Kondo, Miyuki; Krishnan, Srinath; Lincoln, Sara; Lipp, Julius; Mangelsdorf, Kai; Martínez-García, Alfredo; Ménot, Guillemette; Mets, Anchelique; Mollenhauer, Gesine; Ohkouchi, Naohiko; Ossebaar, Jort; Pagani, Mark; Pancost, Richard D.; Pearson, Emma J.; Peterse, Francien; Reichart, Gert-Jan; Schaeffer, Philippe; Schmitt, Gaby; Schwark, Lorenz; Shah, Sunita R.; Smith, Richard W.; Smittenberg, Rienk H.; Summons, Roger E.; Takano, Yoshinori; Talbot, Helen M.; Taylor, Kyle W. R.; Tarozo, Rafael; Uchida, Masao; van Dongen, Bart E.; Van Mooy, Benjamin A. S.; Wang, Jinxiang; Warren, Courtney; Weijers, Johan W. H.; Werne, Josef P.; Woltering, Martijn; Xie, Shucheng; Yamamoto, Masanobu; Yang, Huan; Zhang, Chuanlun L.; Zhang, Yige; Zhao, Meixun; Damsté, Jaap S. Sinninghe

2013-12-01

Two commonly used proxies based on the distribution of glycerol dialkyl glycerol tetraethers (GDGTs) are the TEX86 (TetraEther indeX of 86 carbon atoms) paleothermometer for sea surface temperature reconstructions and the BIT (Branched Isoprenoid Tetraether) index for reconstructing soil organic matter input to the ocean. An initial round-robin study of two sediment extracts, in which 15 laboratories participated, showed relatively consistent TEX86 values (reproducibility ±3-4°C when translated to temperature) but a large spread in BIT measurements (reproducibility ±0.41 on a scale of 0-1). Here we report results of a second round-robin study with 35 laboratories in which three sediments, one sediment extract, and two mixtures of pure, isolated GDGTs were analyzed. The results for TEX86 and BIT index showed improvement compared to the previous round-robin study. The reproducibility, indicating interlaboratory variation, of TEX86 values ranged from 1.3 to 3.0°C when translated to temperature. These results are similar to those of other temperature proxies used in paleoceanography. Comparison of the results obtained from one of the three sediments showed that TEX86 and BIT indices are not significantly affected by interlaboratory differences in sediment extraction techniques. BIT values of the sediments and extracts were at the extremes of the index with values close to 0 or 1, and showed good reproducibility (ranging from 0.013 to 0.042). However, the measured BIT values for the two GDGT mixtures, with known molar ratios of crenarchaeol and branched GDGTs, had intermediate BIT values and showed poor reproducibility and a large overestimation of the "true" (i.e., molar-based) BIT index. The latter is likely due to, among other factors, the higher mass spectrometric response of branched GDGTs compared to crenarchaeol, which also varies among mass spectrometers. Correction for this different mass spectrometric response showed a considerable improvement in the reproducibility of BIT index measurements among laboratories, as well as a substantially improved estimation of molar-based BIT values. This suggests that standard mixtures should be used in order to obtain consistent, and molar-based, BIT values.

MIGS-GPU: Microarray Image Gridding and Segmentation on the GPU.

PubMed

Katsigiannis, Stamos; Zacharia, Eleni; Maroulis, Dimitris

2017-05-01

Complementary DNA (cDNA) microarray is a powerful tool for simultaneously studying the expression level of thousands of genes. Nevertheless, the analysis of microarray images remains an arduous and challenging task due to the poor quality of the images that often suffer from noise, artifacts, and uneven background. In this study, the MIGS-GPU [Microarray Image Gridding and Segmentation on Graphics Processing Unit (GPU)] software for gridding and segmenting microarray images is presented. MIGS-GPU's computations are performed on the GPU by means of the compute unified device architecture (CUDA) in order to achieve fast performance and increase the utilization of available system resources. Evaluation on both real and synthetic cDNA microarray images showed that MIGS-GPU provides better performance than state-of-the-art alternatives, while the proposed GPU implementation achieves significantly lower computational times compared to the respective CPU approaches. Consequently, MIGS-GPU can be an advantageous and useful tool for biomedical laboratories, offering a user-friendly interface that requires minimum input in order to run.

The MicroArray Quality Control (MAQC) project shows inter- and intraplatform reproducibility of gene expression measurements

PubMed Central

2012-01-01

Over the last decade, the introduction of microarray technology has had a profound impact on gene expression research. The publication of studies with dissimilar or altogether contradictory results, obtained using different microarray platforms to analyze identical RNA samples, has raised concerns about the reliability of this technology. The MicroArray Quality Control (MAQC) project was initiated to address these concerns, as well as other performance and data analysis issues. Expression data on four titration pools from two distinct reference RNA samples were generated at multiple test sites using a variety of microarray-based and alternative technology platforms. Here we describe the experimental design and probe mapping efforts behind the MAQC project. We show intraplatform consistency across test sites as well as a high level of interplatform concordance in terms of genes identified as differentially expressed. This study provides a resource that represents an important first step toward establishing a framework for the use of microarrays in clinical and regulatory settings. PMID:16964229

Women's experiences receiving abnormal prenatal chromosomal microarray testing results.

PubMed

Bernhardt, Barbara A; Soucier, Danielle; Hanson, Karen; Savage, Melissa S; Jackson, Laird; Wapner, Ronald J

2013-02-01

Genomic microarrays can detect copy-number variants not detectable by conventional cytogenetics. This technology is diffusing rapidly into prenatal settings even though the clinical implications of many copy-number variants are currently unknown. We conducted a qualitative pilot study to explore the experiences of women receiving abnormal results from prenatal microarray testing performed in a research setting. Participants were a subset of women participating in a multicenter prospective study "Prenatal Cytogenetic Diagnosis by Array-based Copy Number Analysis." Telephone interviews were conducted with 23 women receiving abnormal prenatal microarray results. We found that five key elements dominated the experiences of women who had received abnormal prenatal microarray results: an offer too good to pass up, blindsided by the results, uncertainty and unquantifiable risks, need for support, and toxic knowledge. As prenatal microarray testing is increasingly used, uncertain findings will be common, resulting in greater need for careful pre- and posttest counseling, and more education of and resources for providers so they can adequately support the women who are undergoing testing.

The Glycan Microarray Story from Construction to Applications.

PubMed

Hyun, Ji Young; Pai, Jaeyoung; Shin, Injae

2017-04-18

Not only are glycan-mediated binding processes in cells and organisms essential for a wide range of physiological processes, but they are also implicated in various pathological processes. As a result, elucidation of glycan-associated biomolecular interactions and their consequences is of great importance in basic biological research and biomedical applications. In 2002, we and others were the first to utilize glycan microarrays in efforts aimed at the rapid analysis of glycan-associated recognition events. Because they contain a number of glycans immobilized in a dense and orderly manner on a solid surface, glycan microarrays enable multiple parallel analyses of glycan-protein binding events while utilizing only small amounts of glycan samples. Therefore, this microarray technology has become a leading edge tool in studies aimed at elucidating roles played by glycans and glycan binding proteins in biological systems. In this Account, we summarize our efforts on the construction of glycan microarrays and their applications in studies of glycan-associated interactions. Immobilization strategies of functionalized and unmodified glycans on derivatized glass surfaces are described. Although others have developed immobilization techniques, our efforts have focused on improving the efficiencies and operational simplicity of microarray construction. The microarray-based technology has been most extensively used for rapid analysis of the glycan binding properties of proteins. In addition, glycan microarrays have been employed to determine glycan-protein interactions quantitatively, detect pathogens, and rapidly assess substrate specificities of carbohydrate-processing enzymes. More recently, the microarrays have been employed to identify functional glycans that elicit cell surface lectin-mediated cellular responses. Owing to these efforts, it is now possible to use glycan microarrays to expand the understanding of roles played by glycans and glycan binding proteins in biological systems.

Interlaboratory study for the assessment of potential irritative properties of hygiene products on the hamster cheek pouch.

PubMed

Bourrinet, P; Conduzorgues, J P; Dutertre, H; Macabies, J; Masson, P; Maurin, J; Mercier, O

1995-02-01

An interlaboratory study was carried out to determine the feasibility and reliability of a method using the hamster cheek pouch as a model for assessing the potential irritative properties of substances intended to be applied to the lips or other mucous membranes. The test substances were applied once daily to both pouches for 14 consecutive days. Local and general tolerances were appraised throughout the study. At the end of the study, histologic examination of the pouches and the main organs was performed. Results of the feasibility study, conducted on various types of commercial products, indicated that this model is suitable for preparations of various consistence and composition. Results of the reliability study, carried out on gel-type preparations containing various concentrations of a known irritant, sodium lauryl sulfate, indicated that the method elicits a dose-dependent reaction for this compound. This hamster cheek pouch method was reproducible for the various parameters under consideration: local tolerance, general tolerance, histologic examination. For all products, results were in good agreement among the various laboratories participating in the study. The French regulatory authorities of the Fraud Repression Department have accepted it as an official method for the evaluation of the potential irritative properties of cosmetics and hygiene products intended to be applied to the lips or other mucous membranes.

The MicroArray Quality Control (MAQC) project shows inter- and intraplatform reproducibility of gene expression measurements

EPA Science Inventory

Over the last decade, the introduction of microarray technology has had a profound impact on gene expression research. The publication of studies with dissimilar or altogether contradictory results, obtained using different microarray platforms to analyze identical RNA samples, ...

Flow-pattern Guided Fabrication of High-density Barcode Antibody Microarray

PubMed Central

Ramirez, Lisa S.; Wang, Jun

2016-01-01

Antibody microarray as a well-developed technology is currently challenged by a few other established or emerging high-throughput technologies. In this report, we renovate the antibody microarray technology by using a novel approach for manufacturing and by introducing new features. The fabrication of our high-density antibody microarray is accomplished through perpendicularly oriented flow-patterning of single stranded DNAs and subsequent conversion mediated by DNA-antibody conjugates. This protocol outlines the critical steps in flow-patterning DNA, producing and purifying DNA-antibody conjugates, and assessing the quality of the fabricated microarray. The uniformity and sensitivity are comparable with conventional microarrays, while our microarray fabrication does not require the assistance of an array printer and can be performed in most research laboratories. The other major advantage is that the size of our microarray units is 10 times smaller than that of printed arrays, offering the unique capability of analyzing functional proteins from single cells when interfacing with generic microchip designs. This barcode technology can be widely employed in biomarker detection, cell signaling studies, tissue engineering, and a variety of clinical applications. PMID:26780370

Transfection microarray and the applications.

PubMed

Miyake, Masato; Yoshikawa, Tomohiro; Fujita, Satoshi; Miyake, Jun

2009-05-01

Microarray transfection has been extensively studied for high-throughput functional analysis of mammalian cells. However, control of efficiency and reproducibility are the critical issues for practical use. By using solid-phase transfection accelerators and nano-scaffold, we provide a highly efficient and reproducible microarray-transfection device, "transfection microarray". The device would be applied to the limited number of available primary cells and stem cells not only for large-scale functional analysis but also reporter-based time-lapse cellular event analysis.

DNA microarrays and their use in dermatology.

PubMed

Mlakar, Vid; Glavac, Damjan

2007-03-01

Multiple different DNA microarray technologies are available on the market today. They can be used for studying either DNA or RNA with the purpose of identifying and explaining the role of genes involved in different processes. This paper reviews different DNA microarray platforms available for such studies and their usage in cases of malignant melanomas, psoriasis, and exposure of keratinocytes and melanocytes to UV illumination.

An interlaboratory study to test instrument performance of hydrogen dual-inlet isotope-ratio mass spectrometers

USGS Publications Warehouse

Brand, Willi A.; Coplen, T.B.

2001-01-01

An interlaboratory comparison of forty isotope-ratio mass spectrometers of different ages from several vendors has been performed to test 2H/1H performance with hydrogen gases of three different isotopic compositions. The isotope-ratio results (unsufficiently corrected for H3+ contribution to the m/z = 3 collector, uncorrected for valve leakage in the change-over valves, etc.) expressed relative to one of these three gases covered a wide range of values: -630??? to -790??? for the second gas and -368??? to -462??? for the third gas. After normalizing the isotopic abundances of these test gases (linearly adjusting the ?? values so that the gases with the lowest and highest 2H content were identical for all laboratories), the standard deviation of the 40 measurements of the intermediate gas was a remarkably low 0.85???. It is concluded that the use of scaling factors is mandatory for providing accurate internationally comparable isotope-abundance values. Linear scaling for the isotope-ratio scales of gaseous hydrogen mass spectrometers is completely adequate. ?? Springer-Verlag 2001.

MICROARRAY QUALITY CONTROL PROJECT: A COMPREHENSIVE GENE EXPRESSION TECHNOLOGY SURVEY DEMONSTRATES MEASURABLE CONSISTENCY AND CONCORDANT RESULTS BETWEEN PLATFORMS

EPA Science Inventory

Over the last decade, the introduction of microarray technology has had a profound impact on gene expression research. The publication of studies with dissimilar or altogether contradictory results, obtained using different microarray platforms to analyze identical RNA samples, h...

Applicability of the DPPH assay for evaluating the antioxidant capacity of food additives - inter-laboratory evaluation study -.

PubMed

Shimamura, Tomoko; Sumikura, Yoshihiro; Yamazaki, Takeshi; Tada, Atsuko; Kashiwagi, Takehiro; Ishikawa, Hiroya; Matsui, Toshiro; Sugimoto, Naoki; Akiyama, Hiroshi; Ukeda, Hiroyuki

2014-01-01

An inter-laboratory evaluation study was conducted in order to evaluate the antioxidant capacity of food additives by using a 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay. Four antioxidants used as existing food additives (i.e., tea extract, grape seed extract, enju extract, and d-α-tocopherol) and 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox) were used as analytical samples, and 14 laboratories participated in this study. The repeatability relative standard deviation (RSD(r)) of the IC50 of Trolox, four antioxidants, and the Trolox equivalent antioxidant capacity (TEAC) were 1.8-2.2%, 2.2-2.9%, and 2.1-2.5%, respectively. Thus, the proposed DPPH assay showed good performance within the same laboratory. The reproducibility relative standard deviation (RSD(R)) of IC50 of Trolox, four antioxidants, and TEAC were 4.0-7.9%, 6.0-11%, and 3.7-9.3%, respectively. The RSD(R)/RSD(r) values of TEAC were lower than, or nearly equal to, those of IC50 of the four antioxidants, suggesting that the use of TEAC was effective for reducing the variance among the laboratories. These results showed that the proposed DPPH assay could be used as a standard method to evaluate the antioxidant capacity of food additives.

Predictive performance and inter-laboratory reproducibility in assessing eye irritation potential of water- and oil-soluble mixtures using the Short Time Exposure test method.

PubMed

Abo, Takayuki; Hilberer, Allison; Behle-Wagner, Christine; Watanabe, Mika; Cameron, David; Kirst, Annette; Nukada, Yuko; Yuki, Takuo; Araki, Daisuke; Sakaguchi, Hitoshi; Itagaki, Hiroshi

2018-04-01

The Short Time Exposure (STE) test method is an alternative method for assessing eye irritation potential using Statens Seruminstitut Rabbit Cornea cells and has been adopted as test guideline 491 by the Organisation for Economic Co-operation and Development. Its good predictive performance in identifying the Globally Harmonized System (GHS) No Category (NC) or Irritant Category has been demonstrated in evaluations of water-soluble substances, oil-soluble substances, and water-soluble mixtures. However, the predictive performance for oil-soluble mixtures was not evaluated. Twenty-four oil-soluble mixtures were evaluated using the STE test method. The GHS NC or Irritant Category of 22 oil-soluble mixtures were consistent with that of a Reconstructed human Cornea-like Epithelium (RhCE) test method. Inter-laboratory reproducibility was then confirmed using 20 water- and oil-soluble mixtures blind-coded. The concordance in GHS NC or Irritant Category among four laboratories was 90%-100%. In conclusion, the concordance in comparison with the results of RhCE test method using 24 oil-soluble mixtures and inter-laboratory reproducibility using 20 water- and oil-soluble mixtures blind-coded were good, indicating that the STE test method is a suitable alternative for predicting the eye irritation potential of both substances and mixtures. Copyright © 2018 Elsevier Ltd. All rights reserved.

First Interlaboratory Comparison on Calibration of Temperature-Controlled Enclosures in Turkey

NASA Astrophysics Data System (ADS)

Uytun, A.; Kalemci, M.

2017-11-01

The number of accredited laboratories in the field of calibration of temperature-controlled enclosures has been increasing in Turkey. One of the main criteria demonstrating the competence of a calibration laboratory is successful participation in interlaboratory comparisons. Therefore, TUBITAK UME Temperature Laboratory organized the first interlaboratory comparison on "Calibration of Temperature-Controlled Enclosures" in Turkey as a pilot laboratory between January and November, 2013. Forty accredited laboratories which provide routine calibration services to the industry in this field participated in the comparison. The standards used during the comparison was a climatic chamber for the measurements at -40 {°}C, -20 {°}C, 40 {°}C and 100 {°}C and an oven for the measurements at 200 {°}C. The protocol of the comparison was prepared considering guide EURAMET cg-20 and BS EN/IEC standards 600068-3-5 and 600068-3-11. During the comparison measurements, each participant had the liberty to choose the most convenient calibration points in terms of their accreditation scope among the values mentioned above and carried out on-site measurements at UME. The details and the results of this comparison are given in the paper. Determination of the statistical consistency of the results with the uncertainties given by the participants can be assessed by the method of En value assessment for each laboratory. En values for all measurement results based on the results of pilot and participating laboratories were calculated.

Design of microarray experiments for genetical genomics studies.

PubMed

Bueno Filho, Júlio S S; Gilmour, Steven G; Rosa, Guilherme J M

2006-10-01

Microarray experiments have been used recently in genetical genomics studies, as an additional tool to understand the genetic mechanisms governing variation in complex traits, such as for estimating heritabilities of mRNA transcript abundances, for mapping expression quantitative trait loci, and for inferring regulatory networks controlling gene expression. Several articles on the design of microarray experiments discuss situations in which treatment effects are assumed fixed and without any structure. In the case of two-color microarray platforms, several authors have studied reference and circular designs. Here, we discuss the optimal design of microarray experiments whose goals refer to specific genetic questions. Some examples are used to illustrate the choice of a design for comparing fixed, structured treatments, such as genotypic groups. Experiments targeting single genes or chromosomic regions (such as with transgene research) or multiple epistatic loci (such as within a selective phenotyping context) are discussed. In addition, microarray experiments in which treatments refer to families or to subjects (within family structures or complex pedigrees) are presented. In these cases treatments are more appropriately considered to be random effects, with specific covariance structures, in which the genetic goals relate to the estimation of genetic variances and the heritability of transcriptional abundances.

SIMULATION AND VISUALIZATION OF FLOW PATTERN IN MICROARRAYS FOR LIQUID PHASE OLIGONUCLEOTIDE AND PEPTIDE SYNTHESIS

PubMed Central

O-Charoen, Sirimon; Srivannavit, Onnop; Gulari, Erdogan

2008-01-01

Microfluidic microarrays have been developed for economical and rapid parallel synthesis of oligonucleotide and peptide libraries. For a synthesis system to be reproducible and uniform, it is crucial to have a uniform reagent delivery throughout the system. Computational fluid dynamics (CFD) is used to model and simulate the microfluidic microarrays to study geometrical effects on flow patterns. By proper design geometry, flow uniformity could be obtained in every microreactor in the microarrays. PMID:17480053

The application of DNA microarrays in gene expression analysis.

PubMed

van Hal, N L; Vorst, O; van Houwelingen, A M; Kok, E J; Peijnenburg, A; Aharoni, A; van Tunen, A J; Keijer, J

2000-03-31

DNA microarray technology is a new and powerful technology that will substantially increase the speed of molecular biological research. This paper gives a survey of DNA microarray technology and its use in gene expression studies. The technical aspects and their potential improvements are discussed. These comprise array manufacturing and design, array hybridisation, scanning, and data handling. Furthermore, it is discussed how DNA microarrays can be applied in the working fields of: safety, functionality and health of food and gene discovery and pathway engineering in plants.

Fully Automated Complementary DNA Microarray Segmentation using a Novel Fuzzy-based Algorithm.

PubMed

Saberkari, Hamidreza; Bahrami, Sheyda; Shamsi, Mousa; Amoshahy, Mohammad Javad; Ghavifekr, Habib Badri; Sedaaghi, Mohammad Hossein

2015-01-01

DNA microarray is a powerful approach to study simultaneously, the expression of 1000 of genes in a single experiment. The average value of the fluorescent intensity could be calculated in a microarray experiment. The calculated intensity values are very close in amount to the levels of expression of a particular gene. However, determining the appropriate position of every spot in microarray images is a main challenge, which leads to the accurate classification of normal and abnormal (cancer) cells. In this paper, first a preprocessing approach is performed to eliminate the noise and artifacts available in microarray cells using the nonlinear anisotropic diffusion filtering method. Then, the coordinate center of each spot is positioned utilizing the mathematical morphology operations. Finally, the position of each spot is exactly determined through applying a novel hybrid model based on the principle component analysis and the spatial fuzzy c-means clustering (SFCM) algorithm. Using a Gaussian kernel in SFCM algorithm will lead to improving the quality in complementary DNA microarray segmentation. The performance of the proposed algorithm has been evaluated on the real microarray images, which is available in Stanford Microarray Databases. Results illustrate that the accuracy of microarray cells segmentation in the proposed algorithm reaches to 100% and 98% for noiseless/noisy cells, respectively.

Implementation of a National Reference Laboratory for Buruli Ulcer Disease in Togo

PubMed Central

Badziklou, Kossi; Halatoko, Wemboo Afiwa; Maman, Issaka; Vogel, Felix; Bidjada, Bawimodom; Awoussi, Koffi Somenou; Piten, Ebekalisai; Helfrich, Kerstin; Mengele, Carolin; Nitschke, Jörg; Amekuse, Komi; Wiedemann, Franz Xaver; Diefenhardt, Adolf; Kobara, Basile; Herbinger, Karl–Heinz; Kere, Abiba Banla; Prince-David, Mireille; Löscher, Thomas; Bretzel, Gisela

2013-01-01

Background In a previous study PCR analysis of clinical samples from suspected cases of Buruli ulcer disease (BUD) from Togo and external quality assurance (EQA) for local microscopy were conducted at an external reference laboratory in Germany. The relatively poor performance of local microscopy as well as effort and time associated with shipment of PCR samples necessitated the implementation of stringent EQA measures and availability of local laboratory capacity. This study describes the approach to implementation of a national BUD reference laboratory in Togo. Methodology Large scale outreach activities accompanied by regular training programs for health care professionals were conducted in the regions “Maritime” and “Central,” standard operating procedures defined all processes in participating laboratories (regional, national and external reference laboratories) as well as the interaction between laboratories and partners in the field. Microscopy was conducted at regional level and slides were subjected to EQA at national and external reference laboratories. For PCR analysis, sample pairs were collected and subjected to a dry-reagent-based IS2404-PCR (DRB-PCR) at national level and standard IS2404 PCR followed by IS2404 qPCR analysis of negative samples at the external reference laboratory. Principal Findings The inter-laboratory concordance rates for microscopy ranged from 89% to 94%; overall, microscopy confirmed 50% of all suspected BUD cases. The inter-laboratory concordance rate for PCR was 96% with an overall PCR case confirmation rate of 78%. Compared to a previous study, the rate of BUD patients with non-ulcerative lesions increased from 37% to 50%, the mean duration of disease before clinical diagnosis decreased significantly from 182.6 to 82.1 days among patients with ulcerative lesions, and the percentage of category III lesions decreased from 30.3% to 19.2%. Conclusions High inter-laboratory concordance rates as well as case confirmation rates of 50% (microscopy), 71% (PCR at national level), and 78% (including qPCR confirmation at external reference laboratory) suggest high standards of BUD diagnostics. The increase of non-ulcerative lesions, as well as the decrease in diagnostic delay and category III lesions, prove the effect of comprehensive EQA and training measures involving also procedures outside the laboratory. PMID:23359828

Implementation of a national reference laboratory for Buruli ulcer disease in Togo.

PubMed

Beissner, Marcus; Huber, Kristina Lydia; Badziklou, Kossi; Halatoko, Wemboo Afiwa; Maman, Issaka; Vogel, Felix; Bidjada, Bawimodom; Awoussi, Koffi Somenou; Piten, Ebekalisai; Helfrich, Kerstin; Mengele, Carolin; Nitschke, Jörg; Amekuse, Komi; Wiedemann, Franz Xaver; Diefenhardt, Adolf; Kobara, Basile; Herbinger, Karl-Heinz; Kere, Abiba Banla; Prince-David, Mireille; Löscher, Thomas; Bretzel, Gisela

2013-01-01

In a previous study PCR analysis of clinical samples from suspected cases of Buruli ulcer disease (BUD) from Togo and external quality assurance (EQA) for local microscopy were conducted at an external reference laboratory in Germany. The relatively poor performance of local microscopy as well as effort and time associated with shipment of PCR samples necessitated the implementation of stringent EQA measures and availability of local laboratory capacity. This study describes the approach to implementation of a national BUD reference laboratory in Togo. Large scale outreach activities accompanied by regular training programs for health care professionals were conducted in the regions "Maritime" and "Central," standard operating procedures defined all processes in participating laboratories (regional, national and external reference laboratories) as well as the interaction between laboratories and partners in the field. Microscopy was conducted at regional level and slides were subjected to EQA at national and external reference laboratories. For PCR analysis, sample pairs were collected and subjected to a dry-reagent-based IS2404-PCR (DRB-PCR) at national level and standard IS2404 PCR followed by IS2404 qPCR analysis of negative samples at the external reference laboratory. The inter-laboratory concordance rates for microscopy ranged from 89% to 94%; overall, microscopy confirmed 50% of all suspected BUD cases. The inter-laboratory concordance rate for PCR was 96% with an overall PCR case confirmation rate of 78%. Compared to a previous study, the rate of BUD patients with non-ulcerative lesions increased from 37% to 50%, the mean duration of disease before clinical diagnosis decreased significantly from 182.6 to 82.1 days among patients with ulcerative lesions, and the percentage of category III lesions decreased from 30.3% to 19.2%. High inter-laboratory concordance rates as well as case confirmation rates of 50% (microscopy), 71% (PCR at national level), and 78% (including qPCR confirmation at external reference laboratory) suggest high standards of BUD diagnostics. The increase of non-ulcerative lesions, as well as the decrease in diagnostic delay and category III lesions, prove the effect of comprehensive EQA and training measures involving also procedures outside the laboratory.

Use of HOMA-IR to diagnose non-alcoholic fatty liver disease: a population-based and inter-laboratory study.

PubMed

Isokuortti, Elina; Zhou, You; Peltonen, Markku; Bugianesi, Elisabetta; Clement, Karine; Bonnefont-Rousselot, Dominique; Lacorte, Jean-Marc; Gastaldelli, Amalia; Schuppan, Detlef; Schattenberg, Jörn M; Hakkarainen, Antti; Lundbom, Nina; Jousilahti, Pekka; Männistö, Satu; Keinänen-Kiukaanniemi, Sirkka; Saltevo, Juha; Anstee, Quentin M; Yki-Järvinen, Hannele

2017-10-01

Recent European guidelines for non-alcoholic fatty liver disease (NAFLD) call for reference values for HOMA-IR. In this study, we aimed to determine: (1) the upper limit of normal HOMA-IR in two population-based cohorts; (2) the HOMA-IR corresponding to NAFLD; (3) the effect of sex and PNPLA3 genotype at rs738409 on HOMA-IR; and (4) inter-laboratory variations in HOMA-IR. We identified healthy individuals in two population-based cohorts (FINRISK 2007 [n = 5024] and the Programme for Prevention of Type 2 Diabetes in Finland [FIN-D2D; n = 2849]) to define the upper 95th percentile of HOMA-IR. Non-obese individuals with normal fasting glucose levels, no excessive alcohol use, no known diseases and no use of any drugs were considered healthy. The optimal HOMA-IR cut-off for NAFLD (liver fat ≥5.56%, based on the Dallas Heart Study) was determined in 368 non-diabetic individuals (35% with NAFLD), whose liver fat was measured using proton magnetic resonance spectroscopy ( 1 H-MRS). Samples from ten individuals were simultaneously analysed for HOMA-IR in seven European laboratories. The upper 95th percentiles of HOMA-IR were 1.9 and 2.0 in healthy individuals in the FINRISK (n = 1167) and FIN-D2D (n = 459) cohorts. Sex or PNPLA3 genotype did not influence these values. The optimal HOMA-IR cut-off for NAFLD was 1.9 (sensitivity 87%, specificity 79%). A HOMA-IR of 2.0 corresponded to normal liver fat (<5.56% on 1 H-MRS) in linear regression analysis. The 2.0 HOMA-IR measured in Helsinki corresponded to 1.3, 1.6, 1.8, 1.8, 2.0 and 2.1 in six other laboratories. The inter-laboratory CV% of HOMA-IR was 25% due to inter-assay variation in insulin (25%) rather than glucose (5%) measurements. The upper limit of HOMA-IR in population-based cohorts closely corresponds to that of normal liver fat. Standardisation of insulin assays would be the first step towards definition of normal values for HOMA-IR.

Direct labeling of serum proteins by fluorescent dye for antibody microarray.

PubMed

Klimushina, M V; Gumanova, N G; Metelskaya, V A

2017-05-06

Analysis of serum proteome by antibody microarray is used to identify novel biomarkers and to study signaling pathways including protein phosphorylation and protein-protein interactions. Labeling of serum proteins is important for optimal performance of the antibody microarray. Proper choice of fluorescent label and optimal concentration of protein loaded on the microarray ensure good quality of imaging that can be reliably scanned and processed by the software. We have optimized direct serum protein labeling using fluorescent dye Arrayit Green 540 (Arrayit Corporation, USA) for antibody microarray. Optimized procedure produces high quality images that can be readily scanned and used for statistical analysis of protein composition of the serum. Copyright © 2017 Elsevier Inc. All rights reserved.

Interlaboratory comparison of mineral constituents in a sample from the Herrin (No. 6) coal bed from Illinois

USGS Publications Warehouse

Finkelman, Robert B.; Fiene, F.L.; Miller, R.N.; Simon, F.O.

1984-01-01

Approximately 20 kg of the Herrin (No. 6) coal was collected from a strip mine in St. Clair County, Ill. A 10-kg portion was ground to -60 mesh, homogenized, and riffled into 128 splits of 70-80 g each. Homogeneity of these splits was confirmed by moisture, ash, and sulfur analyses of six randomly selected splits. Results of these analyses were within the ASTM (American Society for Testing and Materials) guidelines for interlaboratory precision. Splits of the Herrin (No. 6) coal were then transmitted to more than 30 laboratories for analysis. Low-temperature plasma oxidation was used to isolate inorganic matter for quantitative chemical and mineralogical analysis. Despite a wide variation in ashing conditions, only minor variations in ash yields were obtained; these variations were attributed to differences in operating temperature and moisture content. Mineralogical analyses of low-temperature ash (LTA) concentrates prepared by five different laboratories indicated variations within the limits of analytical error. The mean values, in weight percent, for the major minerals are as follows: calcite, 9; quartz, 20; pyrite, 23; kaolinite, 14; and illite+mixed-layer clays, 31. Normative mineralogical calculations and Fourier transform infrared analysis (FTIR) yielded results similar to those obtained from X-ray diffraction (XRD). Choosing appropriate mineral standards was found to be critical for the proper use of analytical techniques such as XRD and FTIR. Good interlaboratory agreement was obtained for most major, minor, and trace elements despite differences in analytical procedures and in the type of sample analyzed (coal, high-temperature ash, or LTA). Discrepancies between analyses for zinc, strontium, manganese, and iron may be attributed to sampling inhomogeneity problems. Mossbauer spectroscopy showed that approximately 44 percent of the pyritic sulfur was lost through weathering in the first year after preparation of the interlaboratory sample. Szomolnokite and possibly coquimbite and jarosite were also identified. Scanning electron microscopy studies indicated ubiquitous pyrite framboids and, less commonly, euhedral crystals, skeletal grains, irregularly shaped particles, and vein fillings. Minor accessory minerals such as rare-earth phosphates and possibly silicates, zircon, barium sulfate, titanium oxide, and sphalerite were also found. The textural evidence indicates that the minerals in the banded material are detrital whereas the minerals occurring as vein and pore fillings are authigenic. Magnetic measurements indicate that coal crushed in a steel pulverizer is contaminated by small quantities of abrasion fragments from the crusher, which seriously affect the measured magnetic properties of the coal.

Comparison of RNA-seq and microarray-based models for clinical endpoint prediction.

PubMed

Zhang, Wenqian; Yu, Ying; Hertwig, Falk; Thierry-Mieg, Jean; Zhang, Wenwei; Thierry-Mieg, Danielle; Wang, Jian; Furlanello, Cesare; Devanarayan, Viswanath; Cheng, Jie; Deng, Youping; Hero, Barbara; Hong, Huixiao; Jia, Meiwen; Li, Li; Lin, Simon M; Nikolsky, Yuri; Oberthuer, André; Qing, Tao; Su, Zhenqiang; Volland, Ruth; Wang, Charles; Wang, May D; Ai, Junmei; Albanese, Davide; Asgharzadeh, Shahab; Avigad, Smadar; Bao, Wenjun; Bessarabova, Marina; Brilliant, Murray H; Brors, Benedikt; Chierici, Marco; Chu, Tzu-Ming; Zhang, Jibin; Grundy, Richard G; He, Min Max; Hebbring, Scott; Kaufman, Howard L; Lababidi, Samir; Lancashire, Lee J; Li, Yan; Lu, Xin X; Luo, Heng; Ma, Xiwen; Ning, Baitang; Noguera, Rosa; Peifer, Martin; Phan, John H; Roels, Frederik; Rosswog, Carolina; Shao, Susan; Shen, Jie; Theissen, Jessica; Tonini, Gian Paolo; Vandesompele, Jo; Wu, Po-Yen; Xiao, Wenzhong; Xu, Joshua; Xu, Weihong; Xuan, Jiekun; Yang, Yong; Ye, Zhan; Dong, Zirui; Zhang, Ke K; Yin, Ye; Zhao, Chen; Zheng, Yuanting; Wolfinger, Russell D; Shi, Tieliu; Malkas, Linda H; Berthold, Frank; Wang, Jun; Tong, Weida; Shi, Leming; Peng, Zhiyu; Fischer, Matthias

2015-06-25

Gene expression profiling is being widely applied in cancer research to identify biomarkers for clinical endpoint prediction. Since RNA-seq provides a powerful tool for transcriptome-based applications beyond the limitations of microarrays, we sought to systematically evaluate the performance of RNA-seq-based and microarray-based classifiers in this MAQC-III/SEQC study for clinical endpoint prediction using neuroblastoma as a model. We generate gene expression profiles from 498 primary neuroblastomas using both RNA-seq and 44 k microarrays. Characterization of the neuroblastoma transcriptome by RNA-seq reveals that more than 48,000 genes and 200,000 transcripts are being expressed in this malignancy. We also find that RNA-seq provides much more detailed information on specific transcript expression patterns in clinico-genetic neuroblastoma subgroups than microarrays. To systematically compare the power of RNA-seq and microarray-based models in predicting clinical endpoints, we divide the cohort randomly into training and validation sets and develop 360 predictive models on six clinical endpoints of varying predictability. Evaluation of factors potentially affecting model performances reveals that prediction accuracies are most strongly influenced by the nature of the clinical endpoint, whereas technological platforms (RNA-seq vs. microarrays), RNA-seq data analysis pipelines, and feature levels (gene vs. transcript vs. exon-junction level) do not significantly affect performances of the models. We demonstrate that RNA-seq outperforms microarrays in determining the transcriptomic characteristics of cancer, while RNA-seq and microarray-based models perform similarly in clinical endpoint prediction. Our findings may be valuable to guide future studies on the development of gene expression-based predictive models and their implementation in clinical practice.

ArrayNinja: An Open Source Platform for Unified Planning and Analysis of Microarray Experiments.

PubMed

Dickson, B M; Cornett, E M; Ramjan, Z; Rothbart, S B

2016-01-01

Microarray-based proteomic platforms have emerged as valuable tools for studying various aspects of protein function, particularly in the field of chromatin biochemistry. Microarray technology itself is largely unrestricted in regard to printable material and platform design, and efficient multidimensional optimization of assay parameters requires fluidity in the design and analysis of custom print layouts. This motivates the need for streamlined software infrastructure that facilitates the combined planning and analysis of custom microarray experiments. To this end, we have developed ArrayNinja as a portable, open source, and interactive application that unifies the planning and visualization of microarray experiments and provides maximum flexibility to end users. Array experiments can be planned, stored to a private database, and merged with the imaged results for a level of data interaction and centralization that is not currently attainable with available microarray informatics tools. © 2016 Elsevier Inc. All rights reserved.

PSU/WES Interlaboratory Comparative Methodology Study of an Experimental Cementitious Repository Seal Material. Report 2. Final Results.

DTIC Science & Technology

1982-03-01

meter 25 11.0 Microstructure by SEM 11.1 Introduction In order to correlate observed physical and mechanical properties in cured grout samples, a...studied at the two laboratories has proper physical properties , phase composi- tions, and microstructures for the materials used and ages covered...Scanning Electron Microscope Resolution Test Specimen ( Al -W) D. B. Ballard Research Material 100 SEM Resolution Test Specimen (AI-W)., is an alloy of

Moving Toward Integrating Gene Expression Profiling into High-throughput Testing:A Gene Expression Biomarker Accurately Predicts Estrogen Receptor α Modulation in a Microarray Compendium

EPA Science Inventory

Microarray profiling of chemical-induced effects is being increasingly used in medium and high-throughput formats. In this study, we describe computational methods to identify molecular targets from whole-genome microarray data using as an example the estrogen receptor α (ERα), ...

Interlaboratory Reproducibility of Droplet Digital Polymerase Chain Reaction Using a New DNA Reference Material Format.

PubMed

Pinheiro, Leonardo B; O'Brien, Helen; Druce, Julian; Do, Hongdo; Kay, Pippa; Daniels, Marissa; You, Jingjing; Burke, Daniel; Griffiths, Kate; Emslie, Kerry R

2017-11-07

Use of droplet digital PCR technology (ddPCR) is expanding rapidly in the diversity of applications and number of users around the world. Access to relatively simple and affordable commercial ddPCR technology has attracted wide interest in use of this technology as a molecular diagnostic tool. For ddPCR to effectively transition to a molecular diagnostic setting requires processes for method validation and verification and demonstration of reproducible instrument performance. In this study, we describe the development and characterization of a DNA reference material (NMI NA008 High GC reference material) comprising a challenging methylated GC-rich DNA template under a novel 96-well microplate format. A scalable process using high precision acoustic dispensing technology was validated to produce the DNA reference material with a certified reference value expressed in amount of DNA molecules per well. An interlaboratory study, conducted using blinded NA008 High GC reference material to assess reproducibility among seven independent laboratories demonstrated less than 4.5% reproducibility relative standard deviation. With the exclusion of one laboratory, laboratories had appropriate technical competency, fully functional instrumentation, and suitable reagents to perform accurate ddPCR based DNA quantification measurements at the time of the study. The study results confirmed that NA008 High GC reference material is fit for the purpose of being used for quality control of ddPCR systems, consumables, instrumentation, and workflow.

Interlaboratory Study of Quality Control Isolates for a Broth Microdilution Method (Modified CLSI M38-A) for Testing Susceptibilities of Dermatophytes to Antifungals▿

PubMed Central

Ghannoum, M. A.; Arthington-Skaggs, B.; Chaturvedi, V.; Espinel-Ingroff, A.; Pfaller, M. A.; Rennie, R.; Rinaldi, M. G.; Walsh, T. J.

2006-01-01

The Clinical and Laboratory Standards Institute (CLSI; formerly National Committee for Clinical Laboratory Standards, or NCCLS) M38-A standard for the susceptibility testing of filamentous fungi does not specifically address the testing of dermatophytes. In 2003, a multicenter study investigated the reproducibility of the microdilution method developed at the Center for Medical Mycology, Cleveland, Ohio, for testing the susceptibility of dermatophytes. Data from that study supported the introduction of this method for testing dermatophytes in the future version of the CLSI M38-A standard. In order for the method to be accepted by CLSI, appropriate quality control isolates needed to be identified. To that end, an interlaboratory study, involving the original six laboratories plus two additional sites, was conducted to evaluate potential candidates for quality control isolates. These candidate strains included five Trichophyton rubrum strains known to have elevated MICs to terbinafine and five Trichophyton mentagrophytes strains. Antifungal agents tested included ciclopirox, fluconazole, griseofulvin, itraconazole, posaconazole, terbinafine, and voriconazole. Based on the data generated, two quality control isolates, one T. rubrum isolate and one T. mentagrophytes isolate, were identified and submitted to the American Type Culture Collection (ATCC) for inclusion as reference strains. Ranges encompassing 95.2 to 97.9% of all data points for all seven drugs were established. PMID:17050812

The Development of Protein Microarrays and Their Applications in DNA-Protein and Protein-Protein Interaction Analyses of Arabidopsis Transcription Factors

PubMed Central

Gong, Wei; He, Kun; Covington, Mike; Dinesh-Kumar, S. P.; Snyder, Michael; Harmer, Stacey L.; Zhu, Yu-Xian; Deng, Xing Wang

2009-01-01

We used our collection of Arabidopsis transcription factor (TF) ORFeome clones to construct protein microarrays containing as many as 802 TF proteins. These protein microarrays were used for both protein-DNA and protein-protein interaction analyses. For protein-DNA interaction studies, we examined AP2/ERF family TFs and their cognate cis-elements. By careful comparison of the DNA-binding specificity of 13 TFs on the protein microarray with previous non-microarray data, we showed that protein microarrays provide an efficient and high throughput tool for genome-wide analysis of TF-DNA interactions. This microarray protein-DNA interaction analysis allowed us to derive a comprehensive view of DNA-binding profiles of AP2/ERF family proteins in Arabidopsis. It also revealed four TFs that bound the EE (evening element) and had the expected phased gene expression under clock-regulation, thus providing a basis for further functional analysis of their roles in clock regulation of gene expression. We also developed procedures for detecting protein interactions using this TF protein microarray and discovered four novel partners that interact with HY5, which can be validated by yeast two-hybrid assays. Thus, plant TF protein microarrays offer an attractive high-throughput alternative to traditional techniques for TF functional characterization on a global scale. PMID:19802365

Use of the Cultex® Radial Flow System as an in vitro exposure method to assess acute pulmonary toxicity of fine dusts and nanoparticles with special focus on the intra- and inter-laboratory reproducibility.

PubMed

Steinritz, Dirk; Möhle, Niklas; Pohl, Christine; Papritz, Mirko; Stenger, Bernhard; Schmidt, Annette; Kirkpatrick, Charles James; Thiermann, Horst; Vogel, Richard; Hoffmann, Sebastian; Aufderheide, Michaela

2013-12-05

Exposure of the respiratory tract to airborne particles (including metal-dusts and nano-particles) is considered as a serious health hazard. For a wide range of substances basic knowledge about the toxic properties and the underlying pathomechanisms is lacking or even completely missing. Legislation demands the toxicological characterization of all chemicals placed on the market until 2018 (REACH). As toxicological in vivo data are rare with regard to acute lung toxicity or exhibit distinct limitations (e.g. inter-species differences) and legislation claims the reduction of animal experiments in general ("3R" principle), profound in vitro models have to be established and characterized to meet these requirements. In this paper we characterize a recently introduced advanced in vitro exposure system (Cultex® RFS) showing a great similarity to the physiological in vivo exposure situation for the assessment of acute pulmonary toxicity of airborne materials. Using the Cultex® RFS, human lung epithelial cells (A549 cells) were exposed to different concentrations of airborne metal dusts (nano- and microscale particles) at the air-liquid-interface (ALI). Cell viability (WST-1 assay) as a parameter of toxicity was assessed 24h after exposure with special focus on the intra- and inter-laboratory (three independent laboratories) reproducibility. Our results show the general applicability of the Cultex® RFS with regard to the requirements of the ECVAM (European Centre for the Validation of Alternative Methods) principles on test validity underlining its robustness and stability. Intra- and inter-laboratory reproducibility can be considered as sufficient if predefined quality criteria are respected. Special attention must be paid to the pure air controls that turned out to be a critical parameter for a rational interpretation of the results. Our results are encouraging and future work is planned to improve the inter-laboratory reproducibility, to consolidate the results so far and to develop a valid prediction model. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Quantifying inter-laboratory variability in stable isotope analysis of ancient skeletal remains.

PubMed

Pestle, William J; Crowley, Brooke E; Weirauch, Matthew T

2014-01-01

Over the past forty years, stable isotope analysis of bone (and tooth) collagen and hydroxyapatite has become a mainstay of archaeological and paleoanthropological reconstructions of paleodiet and paleoenvironment. Despite this method's frequent use across anthropological subdisciplines (and beyond), the present work represents the first attempt at gauging the effects of inter-laboratory variability engendered by differences in a) sample preparation, and b) analysis (instrumentation, working standards, and data calibration). Replicate analyses of a 14C-dated ancient human bone by twenty-one archaeological and paleoecological stable isotope laboratories revealed significant inter-laboratory isotopic variation for both collagen and carbonate. For bone collagen, we found a sizeable range of 1.8‰ for δ13Ccol and 1.9‰ for δ15Ncol among laboratories, but an interpretatively insignificant average pairwise difference of 0.2‰ and 0.4‰ for δ13Ccol and δ15Ncol respectively. For bone hydroxyapatite the observed range increased to a troublingly large 3.5‰ for δ13Cap and 6.7‰ for δ18Oap, with average pairwise differences of 0.6‰ for δ13Cap and a disquieting 2.0‰ for δ18Oap. In order to assess the effects of preparation versus analysis on isotopic variability among laboratories, a subset of the samples prepared by the participating laboratories were analyzed a second time on the same instrument. Based on this duplicate analysis, it was determined that roughly half of the isotopic variability among laboratories could be attributed to differences in sample preparation, with the other half resulting from differences in analysis (instrumentation, working standards, and data calibration). These findings have serious implications for choices made in the preparation and extraction of target biomolecules, the comparison of results obtained from different laboratories, and the interpretation of small differences in bone collagen and hydroxyapatite isotope values. To address the issues arising from inter-laboratory comparisons, we devise a novel measure we term the Minimum Meaningful Difference (MMD), and demonstrate its application.

Quantifying Inter-Laboratory Variability in Stable Isotope Analysis of Ancient Skeletal Remains

PubMed Central

Pestle, William J.; Crowley, Brooke E.; Weirauch, Matthew T.

2014-01-01

Over the past forty years, stable isotope analysis of bone (and tooth) collagen and hydroxyapatite has become a mainstay of archaeological and paleoanthropological reconstructions of paleodiet and paleoenvironment. Despite this method's frequent use across anthropological subdisciplines (and beyond), the present work represents the first attempt at gauging the effects of inter-laboratory variability engendered by differences in a) sample preparation, and b) analysis (instrumentation, working standards, and data calibration). Replicate analyses of a 14C-dated ancient human bone by twenty-one archaeological and paleoecological stable isotope laboratories revealed significant inter-laboratory isotopic variation for both collagen and carbonate. For bone collagen, we found a sizeable range of 1.8‰ for δ13Ccol and 1.9‰ for δ15Ncol among laboratories, but an interpretatively insignificant average pairwise difference of 0.2‰ and 0.4‰ for δ13Ccol and δ15Ncol respectively. For bone hydroxyapatite the observed range increased to a troublingly large 3.5‰ for δ13Cap and 6.7‰ for δ18Oap, with average pairwise differences of 0.6‰ for δ13Cap and a disquieting 2.0‰ for δ18Oap. In order to assess the effects of preparation versus analysis on isotopic variability among laboratories, a subset of the samples prepared by the participating laboratories were analyzed a second time on the same instrument. Based on this duplicate analysis, it was determined that roughly half of the isotopic variability among laboratories could be attributed to differences in sample preparation, with the other half resulting from differences in analysis (instrumentation, working standards, and data calibration). These findings have serious implications for choices made in the preparation and extraction of target biomolecules, the comparison of results obtained from different laboratories, and the interpretation of small differences in bone collagen and hydroxyapatite isotope values. To address the issues arising from inter-laboratory comparisons, we devise a novel measure we term the Minimum Meaningful Difference (MMD), and demonstrate its application. PMID:25061843

Best practices for hybridization design in two-colour microarray analysis.

PubMed

Knapen, Dries; Vergauwen, Lucia; Laukens, Kris; Blust, Ronny

2009-07-01

Two-colour microarrays are a popular platform of choice in gene expression studies. Because two different samples are hybridized on a single microarray, and several microarrays are usually needed in a given experiment, there are many possible ways to combine samples on different microarrays. The actual combination employed is commonly referred to as the 'hybridization design'. Different types of hybridization designs have been developed, all aimed at optimizing the experimental setup for the detection of differentially expressed genes while coping with technical noise. Here, we first provide an overview of the different classes of hybridization designs, discussing their advantages and limitations, and then we illustrate the current trends in the use of different hybridization design types in contemporary research.

Microarray Data Processing Techniques for Genome-Scale Network Inference from Large Public Repositories.

PubMed

Chockalingam, Sriram; Aluru, Maneesha; Aluru, Srinivas

2016-09-19

Pre-processing of microarray data is a well-studied problem. Furthermore, all popular platforms come with their own recommended best practices for differential analysis of genes. However, for genome-scale network inference using microarray data collected from large public repositories, these methods filter out a considerable number of genes. This is primarily due to the effects of aggregating a diverse array of experiments with different technical and biological scenarios. Here we introduce a pre-processing pipeline suitable for inferring genome-scale gene networks from large microarray datasets. We show that partitioning of the available microarray datasets according to biological relevance into tissue- and process-specific categories significantly extends the limits of downstream network construction. We demonstrate the effectiveness of our pre-processing pipeline by inferring genome-scale networks for the model plant Arabidopsis thaliana using two different construction methods and a collection of 11,760 Affymetrix ATH1 microarray chips. Our pre-processing pipeline and the datasets used in this paper are made available at http://alurulab.cc.gatech.edu/microarray-pp.

MALDI-TOF mass spectrometry for quantitative gene expression analysis of acid responses in Staphylococcus aureus.

PubMed

Rode, Tone Mari; Berget, Ingunn; Langsrud, Solveig; Møretrø, Trond; Holck, Askild

2009-07-01

Microorganisms are constantly exposed to new and altered growth conditions, and respond by changing gene expression patterns. Several methods for studying gene expression exist. During the last decade, the analysis of microarrays has been one of the most common approaches applied for large scale gene expression studies. A relatively new method for gene expression analysis is MassARRAY, which combines real competitive-PCR and MALDI-TOF (matrix-assisted laser desorption/ionization time-of-flight) mass spectrometry. In contrast to microarray methods, MassARRAY technology is suitable for analysing a larger number of samples, though for a smaller set of genes. In this study we compare the results from MassARRAY with microarrays on gene expression responses of Staphylococcus aureus exposed to acid stress at pH 4.5. RNA isolated from the same stress experiments was analysed using both the MassARRAY and the microarray methods. The MassARRAY and microarray methods showed good correlation. Both MassARRAY and microarray estimated somewhat lower fold changes compared with quantitative real-time PCR (qRT-PCR). The results confirmed the up-regulation of the urease genes in acidic environments, and also indicated the importance of metal ion regulation. This study shows that the MassARRAY technology is suitable for gene expression analysis in prokaryotes, and has advantages when a set of genes is being analysed for an organism exposed to many different environmental conditions.

Interlaboratory study of DNA extraction from multiple ground samples, multiplex real-time PCR, and multiplex qualitative PCR for individual kernel detection system of genetically modified maize.

PubMed

Akiyama, Hiroshi; Sakata, Kozue; Makiyma, Daiki; Nakamura, Kosuke; Teshima, Reiko; Nakashima, Akie; Ogawa, Asako; Yamagishi, Toru; Futo, Satoshi; Oguchi, Taichi; Mano, Junichi; Kitta, Kazumi

2011-01-01

In many countries, the labeling of grains, feed, and foodstuff is mandatory if the genetically modified (GM) organism content exceeds a certain level of approved GM varieties. We previously developed an individual kernel detection system consisting of grinding individual kernels, DNA extraction from the individually ground kernels, GM detection using multiplex real-time PCR, and GM event detection using multiplex qualitative PCR to analyze the precise commingling level and varieties of GM maize in real sample grains. We performed the interlaboratory study of the DNA extraction with multiple ground samples, multiplex real-time PCR detection, and multiplex qualitative PCR detection to evaluate its applicability, practicality, and ruggedness for the individual kernel detection system of GM maize. DNA extraction with multiple ground samples, multiplex real-time PCR, and multiplex qualitative PCR were evaluated by five laboratories in Japan, and all results from these laboratories were consistent with the expected results in terms of the commingling level and event analysis. Thus, the DNA extraction with multiple ground samples, multiplex real-time PCR, and multiplex qualitative PCR for the individual kernel detection system is applicable and practicable in a laboratory to regulate the commingling level of GM maize grain for GM samples, including stacked GM maize.

Receptor binding assay for paralytic shellfish poisoning toxins: optimization and interlaboratory comparison.

PubMed

Ruberu, Shryamalie R; Liu, Yun-Gang; Wong, Carolyn T; Perera, S Kusum; Langlois, Gregg W; Doucette, Gregory J; Powell, Christine L

2003-01-01

A receptor binding assay (RBA) for detection of paralytic shellfish poisoning (PSP) toxins was formatted for use in a high throughput detection system using microplate scintillation counting. The RBA technology was transferred from the National Ocean Service, which uses a Wallac TriLux 1450 MicroBeta microplate scintillation counter, to the California Department of Health Services, which uses a Packard TopCount scintillation counter. Due to differences in the detector arrangement between these 2 counters, markedly different counting efficiencies were exhibited, requiring optimization of the RBA protocol for the TopCount instrument. Precision, accuracy, and sensitivity [limit of detection = 0.2 microg saxitoxin (STX) equiv/100 g shellfish tissue] of the modified protocol were equivalent to those of the original protocol. The RBA robustness and adaptability were demonstrated by an interlaboratory study, in which STX concentrations in shellfish generated by the TopCount were consistent with MicroBeta-derived values. Comparison of STX reference standards obtained from the U.S. Food and Drug Administration and the National Research Council, Canada, showed no observable differences. This study confirms the RBA's value as a rapid, high throughput screen prior to testing by the conventional mouse bioassay (MBA) and its suitability for providing an early warning of increasing PSP toxicity when toxin levels are below the MBA limit of detection.

An inter-laboratory comparison study on transfer, persistence and recovery of DNA from cable ties.

PubMed

Steensma, Kristy; Ansell, Ricky; Clarisse, Lindy; Connolly, Edward; Kloosterman, Ate D; McKenna, Louise G; van Oorschot, Roland A H; Szkuta, Bianca; Kokshoorn, Bas

2017-11-01

To address questions on the activity that led to the deposition of biological traces in a particular case, general information on the probabilities of transfer, persistence and recovery of cellular material in relevant scenarios is necessary. These figures may be derived from experimental data described in forensic literature when conditions relevant to the case were included. The experimental methodology regarding sampling, DNA extraction, DNA typing and profile interpretation that were used to generate these published data may differ from those applied in the case and thus the applicability of the literature data may be questioned. To assess the level of variability that different laboratories obtain when similar exhibits are analysed, we performed an inter-laboratory study between four partner laboratories. Five sets of 20 cable ties bound by different volunteers were distributed to the participating laboratories and sampled and processed according to the in-house protocols. Differences were found for the amount of retrieved DNA, as well as for the reportability and composition of the DNA profiles. These differences also resulted in different probabilities of transfer, persistence and recovery for each laboratory. Nevertheless, when applied to a case example, these differences resulted in similar assignments of weight of evidence given activity-level propositions. Copyright © 2017 Elsevier B.V. All rights reserved.

A Reference Method for Measuring Emissions of SVOCs in ...

EPA Pesticide Factsheets

Semivolatile organic compounds (SVOCs) are indoor air pollutants that may may have significant adverse effects on human health, and emission of SVOCs from building materials and consumer products is of growing concern. Few chamber studies have been conducted due to the challenges associated with SVOC analysis and the lack of validation procedures. Thus there is an urgent need for a reliable and accurate chamber test method to verify the performance of these measurements. A reference method employing a specially-designed chamber and experimental protocol has been developed and is undergoing extensive evaluation. A pilot interlaboratory study (ILS) has been conducted with five laboratories performing chamber tests under identical conditions. Results showed inter-laboratory variations at 25% for SVOC emission rates, with greater agreement observed between intra-laboratory measurements for most of the participating laboratories. The measured concentration profiles also compared reasonably well to the mechanistic model, demonstrating the feasibility of the proposed reference method to independently assess laboratory performance and validate SVOC emission tests. There is an urgent need for improved understanding of the measurement uncertainties associated with SVOC emissions testing. The creation of specially-designed chambers and well-characterized materials serves as a critical prerequisite for improving the procedure used to measure SVOCs emitted from indoor

Degradation in PV Encapsulation Transmittance: An Interlaboratory Study Toward a Climate-Specific Test

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miller, David C.; Hacke, Peter L.; Kempe, Michael D.

2015-06-14

Reduced optical transmittance of encapsulation resulting from ultraviolet (UV) degradation has frequently been identified as a cause of decreased PV module performance through the life of installations in the field. The present module safety and qualification standards, however, apply short UV doses only capable of examining design robustness or 'infant mortality' failures. Essential information that might be used to screen encapsulation through product lifetime remains unknown. For example, the relative efficacy of xenon-arc and UVA-340 fluorescent sources or the typical range of activation energy for degradation is not quantified. We have conducted an interlaboratory experiment to provide the understanding thatmore » will be used towards developing a climate- and configuration-specific (UV) weathering test. Five representative, known formulations of EVA were studied in addition to one TPU material. Replicate laminated silica/polymer/silica specimens are being examined at 14 institutions using a variety of indoor chambers (including Xe, UVA-340, and metal-halide light sources) or field aging. The solar-weighted transmittance, yellowness index, and the UV cut-off wavelength, determined from the measured hemispherical transmittance, are examined to provide understanding and guidance for the UV light source (lamp type) and temperature used in accelerated UV aging tests. Index Terms -- reliability, durability, thermal activation.« less

Degradation in PV Encapsulation Transmittance: An Interlaboratory Study Towards a Climate-Specific Test: Preprint

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miller, David C.; Annigoni, Eleonora; Ballion, Amal

2015-08-12

Reduced optical transmittance of encapsulants resulting from ultraviolet (UV) degradation has frequently been identified as a cause of decreased PV module performance through the life of service in the field. The present module safety and qualification standards, however, apply short UV doses only capable of examining design robustness or 'infant mortality' failures. Essential information that might be used to screen encapsulation through product lifetime remains unknown. For example, the relative efficacy of xenon-arc and UVA-340 fluorescent sources or the typical range of activation energy for degradation is not quantified. We have conducted an interlaboratory experiment to provide the understanding thatmore » will be used towards developing a climate- and configuration-specific (UV) weathering test. Five representative, known formulations of EVA were studied in addition to one TPU material. Replicate laminated silica/polymer/silica specimens are being examined at 14 institutions using a variety of indoor chambers (including Xenon, UVA-340, and metal-halide light sources) or field aging. The solar-weighted transmittance, yellowness index, and the UV cut-off wavelength, determined from the measured hemispherical transmittance, are examined to provide understanding and guidance for the UV light source (lamp type) and temperature used in accelerated UV aging tests.« less

Degradation in PV Encapsulation Transmittance: An Interlaboratory Study Towards a Climate-Specific Test

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miller, David C.; Annigoni, Eleonora; Ballion, Amal

2015-06-14

Reduced optical transmittance of encapsulants resulting from ultraviolet (UV) degradation has frequently been identified as a cause of decreased PV module performance through the life of service in the field. The present module safety and qualification standards, however, apply short UV doses only capable of examining design robustness or 'infant mortality' failures. Essential information that might be used to screen encapsulation through product lifetime remains unknown. For example, the relative efficacy of xenon-arc and UVA-340 fluorescent sources or the typical range of activation energy for degradation is not quantified. We have conducted an interlaboratory experiment to provide the understanding thatmore » will be used towards developing a climate- and configuration-specific (UV) weathering test. Five representative, known formulations of EVA were studied in addition to one TPU material. Replicate laminated silica/polymer/silica specimens are being examined at 14 institutions using a variety of indoor chambers (including Xenon, UVA-340, and metal-halide light sources) or field aging. The solar-weighted transmittance, yellowness index, and the UV cut-off wavelength, determined from the measured hemispherical transmittance, are examined to provide understanding and guidance for the UV light source (lamp type) and temperature used in accelerated UV aging tests.« less

A Platform for Combined DNA and Protein Microarrays Based on Total Internal Reflection Fluorescence

PubMed Central

Asanov, Alexander; Zepeda, Angélica; Vaca, Luis

2012-01-01

We have developed a novel microarray technology based on total internal reflection fluorescence (TIRF) in combination with DNA and protein bioassays immobilized at the TIRF surface. Unlike conventional microarrays that exhibit reduced signal-to-background ratio, require several stages of incubation, rinsing and stringency control, and measure only end-point results, our TIRF microarray technology provides several orders of magnitude better signal-to-background ratio, performs analysis rapidly in one step, and measures the entire course of association and dissociation kinetics between target DNA and protein molecules and the bioassays. In many practical cases detection of only DNA or protein markers alone does not provide the necessary accuracy for diagnosing a disease or detecting a pathogen. Here we describe TIRF microarrays that detect DNA and protein markers simultaneously, which reduces the probabilities of false responses. Supersensitive and multiplexed TIRF DNA and protein microarray technology may provide a platform for accurate diagnosis or enhanced research studies. Our TIRF microarray system can be mounted on upright or inverted microscopes or interfaced directly with CCD cameras equipped with a single objective, facilitating the development of portable devices. As proof-of-concept we applied TIRF microarrays for detecting molecular markers from Bacillus anthracis, the pathogen responsible for anthrax. PMID:22438738

Variance stabilization and normalization for one-color microarray data using a data-driven multiscale approach.

PubMed

Motakis, E S; Nason, G P; Fryzlewicz, P; Rutter, G A

2006-10-15

Many standard statistical techniques are effective on data that are normally distributed with constant variance. Microarray data typically violate these assumptions since they come from non-Gaussian distributions with a non-trivial mean-variance relationship. Several methods have been proposed that transform microarray data to stabilize variance and draw its distribution towards the Gaussian. Some methods, such as log or generalized log, rely on an underlying model for the data. Others, such as the spread-versus-level plot, do not. We propose an alternative data-driven multiscale approach, called the Data-Driven Haar-Fisz for microarrays (DDHFm) with replicates. DDHFm has the advantage of being 'distribution-free' in the sense that no parametric model for the underlying microarray data is required to be specified or estimated; hence, DDHFm can be applied very generally, not just to microarray data. DDHFm achieves very good variance stabilization of microarray data with replicates and produces transformed intensities that are approximately normally distributed. Simulation studies show that it performs better than other existing methods. Application of DDHFm to real one-color cDNA data validates these results. The R package of the Data-Driven Haar-Fisz transform (DDHFm) for microarrays is available in Bioconductor and CRAN.

A probabilistic framework for microarray data analysis: fundamental probability models and statistical inference.

PubMed

Ogunnaike, Babatunde A; Gelmi, Claudio A; Edwards, Jeremy S

2010-05-21

Gene expression studies generate large quantities of data with the defining characteristic that the number of genes (whose expression profiles are to be determined) exceed the number of available replicates by several orders of magnitude. Standard spot-by-spot analysis still seeks to extract useful information for each gene on the basis of the number of available replicates, and thus plays to the weakness of microarrays. On the other hand, because of the data volume, treating the entire data set as an ensemble, and developing theoretical distributions for these ensembles provides a framework that plays instead to the strength of microarrays. We present theoretical results that under reasonable assumptions, the distribution of microarray intensities follows the Gamma model, with the biological interpretations of the model parameters emerging naturally. We subsequently establish that for each microarray data set, the fractional intensities can be represented as a mixture of Beta densities, and develop a procedure for using these results to draw statistical inference regarding differential gene expression. We illustrate the results with experimental data from gene expression studies on Deinococcus radiodurans following DNA damage using cDNA microarrays. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

Optimal Control of Shock Wave Turbulent Boundary Layer Interactions Using Micro-Array Actuation

NASA Technical Reports Server (NTRS)

Anderson, Bernhard H.; Tinapple, Jon; Surber, Lewis

2006-01-01

The intent of this study on micro-array flow control is to demonstrate the viability and economy of Response Surface Methodology (RSM) to determine optimal designs of micro-array actuation for controlling the shock wave turbulent boundary layer interactions within supersonic inlets and compare these concepts to conventional bleed performance. The term micro-array refers to micro-actuator arrays which have heights of 25 to 40 percent of the undisturbed supersonic boundary layer thickness. This study covers optimal control of shock wave turbulent boundary layer interactions using standard micro-vane, tapered micro-vane, and standard micro-ramp arrays at a free stream Mach number of 2.0. The effectiveness of the three micro-array devices was tested using a shock pressure rise induced by the 10 shock generator, which was sufficiently strong as to separate the turbulent supersonic boundary layer. The overall design purpose of the micro-arrays was to alter the properties of the supersonic boundary layer by introducing a cascade of counter-rotating micro-vortices in the near wall region. In this manner, the impact of the shock wave boundary layer (SWBL) interaction on the main flow field was minimized without boundary bleed.

Inter-laboratory comparison measurements of radiochemical laboratories in Slovakia.

PubMed

Meresová, J; Belanová, A; Vrsková, M

2010-01-01

The first inter-laboratory comparison organized by the radiochemistry laboratory of Water Research Institute (WRI) in Bratislava was carried out in 1993 and since then is it realized on an annual basis and about 10 radiochemical laboratories from all over Slovakia are participating. The gross alpha and gross beta activities, and the activity concentrations of (222)Rn, tritium, and (226)Ra, and U(nat) concentration in synthetic water samples are compared. The distributed samples are covering the concentration range prevailing in potable and surface waters and are prepared by dilution of certified reference materials. Over the course of the years 1993-2008, we observed the improvement in the quality of results for most of the laboratories. However, the success rate of the gross alpha determination activity is not improving as much as the other parameters. Copyright 2009 Elsevier Ltd. All rights reserved.

The determination of ethyl glucuronide in hair: Experiences from nine consecutive interlaboratory comparison rounds.

PubMed

Becker, R; Lô, I; Sporkert, F; Baumgartner, M

2018-07-01

The increasing request for hair ethyl glucuronide (HEtG) in alcohol consumption monitoring according to cut-off levels set by the Society of Hair Testing (SoHT) has triggered a proficiency testing program based on interlaboratory comparisons (ILC). Here, the outcome of nine consecutive ILC rounds organised by the SoHT on the determination of HEtG between 2011 and 2017 is summarised regarding interlaboratory reproducibility and the influence of procedural variants. Test samples prepared from cut hair (1mm) with authentic (in-vivo incorporated) and soaked (in-vitro incorporated) HEtG concentrations up to 80pg/mg were provided for 27-35 participating laboratories. Laboratory results were evaluated according to ISO 5725-5 and provided robust averages and relative reproducibility standard deviations typically between 20 and 35% in reasonable accordance with the prediction of the Horwitz model. Evaluation of results regarding the analytical techniques revealed no significant differences between gas and liquid chromatographic methods In contrast, a detailed evaluation of different sample preparations revealed significantly higher average values in case when pulverised hair is tested compared to cut hair. This observation was reinforced over the different ILC rounds and can be attributed to the increased acceptance and routine of hair pulverisation among laboratories. Further, the reproducibility standard deviations among laboratories performing pulverisation were on average in very good agreement with the prediction of the Horwitz model. Use of sonication showed no effect on the HEtG extraction yield. Copyright © 2018 Elsevier B.V. All rights reserved.

Assessing variability in chemical acute toxicity of unionid mussels: Influence of intra- and inter-laboratory testing, life stage, and species

USGS Publications Warehouse

Raimondo, Sandy; Lilavois, Crystal R.; Lee, Larisa; Augspurger, Tom; Wang, Ning; Ingersoll, Christopher G.; Bauer, Candice R.; Hammer, Edward J.; Barron, Mace G.

2016-01-01

We developed a toxicity database for unionid mussels to examine the extent of intra- and inter-laboratory variability in acute toxicity tests with mussel larvae (glochidia) and juveniles; the extent of differential sensitivity of the two life stages; and the variation in sensitivity among commonly tested mussels (Lampsilis siliquoidea, Utterbackia imbecillis, Villosa iris), commonly tested cladocerans (Daphnia magna, Ceriodaphnia dubia) and fish (Oncorhynchus mykiss, Pimephales promelas, Lepomis macrochirus). The results of these analyses indicate intra-laboratory variability for median effect concentrations (EC50) averaged about 2 fold for both life stages, while inter-laboratory variability averaged 3.6 fold for juvenile mussels and 6.3 fold for glochidia. The EC50s for juveniles and glochidia were within a factor of 2 of each other for 50% of paired records across chemicals, with juveniles more sensitive than glochidia by more than 2 fold for 33% of the comparisons made between life stages. There was a high concurrence of the sensitivity of commonly tested L. siliquoidea, U. imbecillis, and V. iris to that of other mussels. However, this concurrence decreases as the taxonomic distance of the commonly tested cladocerans and fish to mussels increases. The compiled mussel database and determination of data variability will advance risk assessments by including more robust species sensitivity distributions, interspecies correlation estimates, and availability of taxon-specific empirically derived application factors for risk assessment.

A 7-d toxicity test for marine pollutants using the Pacific mysid Mysidopsis intii. 2: Protocol evaluation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Harmon, V.L.; Langdon, C.J.

1996-10-01

The sensitivity of the Pacific coast mysid Mysidopsis intii to pollutants was compared in 7-d toxicity tests with that of the Gulf coast mysid M. bahia and the Pacific coast mysid Holmesimysis costata. Survival and growth responses of M. intii to zinc (maximum acceptable toxicant concentration [MATC] survival and growth, 152 {micro}g/L) were as sensitive as survival of both M. bahia (MATC survival, 152 {micro}g/L) and H. costata (MATC survival, 152 {micro}g/L). In contrast, the 7-d test for M. intii was less sensitive (MATC growth and survival, 4.99 mg/L) than the test for H. costata (MATC survival, 1.99 mg/L) whenmore » sodium dodecyl sulfate (SDS) was used as the toxicant. Interlaboratory evaluation of the 7-d test for M. intii exposed to SDS indicated that the test was reliable. The mean test results for the group of participating laboratories were not significantly different from those of a group of three in-house tests, indicating that shipping and handling did not affect mysid sensitivity to SDS. Mysid growth was not as sensitive to SDS as survival in the interlaboratory tests. Although there were significant differences in median lethal concentration (LC50) values among participating laboratories, coefficients of variation of LC50 and MATC survival values among laboratories were 10.3 and 37%, respectively. These coefficients were comparable to those reported for interlaboratory tests with H. costata.« less

Acid-Soluble Internal Capsules for Closed-Face Cassette Elemental Sampling and Analysis of Workplace Air

PubMed Central

Harper, Martin; Ashley, Kevin

2013-01-01

Airborne particles that are collected using closed-face filter cassettes (CFCs), which are used widely in the sampling of workplace aerosols, can deposit in places other than on the filter and thereby may not be included in the ensuing analysis. A technique for ensuring that internal non-filter deposits are included in the analysis is to collect airborne particles within an acid-soluble internal capsule that, following sampling, can be dissolved along with the filter for subsequent elemental analysis. An interlaboratory study (ILS) was carried out to evaluate the use of cellulosic CFC capsule inserts for their suitability in the determination of trace elements in airborne samples. The ILS was performed in accordance with an applicable ASTM International standard practice, ASTM E691, which describes statistical procedures for investigating interlaboratory precision. Performance evaluation materials consisted of prototype cellulose acetate capsules attached to mixed-cellulose ester filters. Batches of capsules were dosed with Pb-containing materials (standard aqueous solutions, and certified reference material soil and paint). Also, aerosol samples containing nine target analyte elements (As, Cd, Co, Cr, Cu, Fe, Pb, Mn, and Ni) were generated using a multiport sampler; various concentrations and sampling times were employed to yield samples fortified at desired loading levels. Triplicates of spiked capsules at three different loadings were conveyed to each volunteer laboratory; loading levels were unknown to the participants. The laboratories were asked to prepare the samples by acid dissolution and to analyze aliquots of extracted samples by atomic spectrometry in accordance with applicable ASTM International Standards. Participants were asked to report their results in units of μg of each target element per sample. For the elements investigated, interlaboratory precision and recovery estimates from the participating laboratories demonstrated the utility of the cellulosic capsule inserts for the measurement of sampled trace elements. PMID:23548078

Comet assay in reconstructed 3D human epidermal skin models--investigation of intra- and inter-laboratory reproducibility with coded chemicals.

PubMed

Reus, Astrid A; Reisinger, Kerstin; Downs, Thomas R; Carr, Gregory J; Zeller, Andreas; Corvi, Raffaella; Krul, Cyrille A M; Pfuhler, Stefan

2013-11-01

Reconstructed 3D human epidermal skin models are being used increasingly for safety testing of chemicals. Based on EpiDerm™ tissues, an assay was developed in which the tissues were topically exposed to test chemicals for 3h followed by cell isolation and assessment of DNA damage using the comet assay. Inter-laboratory reproducibility of the 3D skin comet assay was initially demonstrated using two model genotoxic carcinogens, methyl methane sulfonate (MMS) and 4-nitroquinoline-n-oxide, and the results showed good concordance among three different laboratories and with in vivo data. In Phase 2 of the project, intra- and inter-laboratory reproducibility was investigated with five coded compounds with different genotoxicity liability tested at three different laboratories. For the genotoxic carcinogens MMS and N-ethyl-N-nitrosourea, all laboratories reported a dose-related and statistically significant increase (P < 0.05) in DNA damage in every experiment. For the genotoxic carcinogen, 2,4-diaminotoluene, the overall result from all laboratories showed a smaller, but significant genotoxic response (P < 0.05). For cyclohexanone (CHN) (non-genotoxic in vitro and in vivo, and non-carcinogenic), an increase compared to the solvent control acetone was observed only in one laboratory. However, the response was not dose related and CHN was judged negative overall, as was p-nitrophenol (p-NP) (genotoxic in vitro but not in vivo and non-carcinogenic), which was the only compound showing clear cytotoxic effects. For p-NP, significant DNA damage generally occurred only at doses that were substantially cytotoxic (>30% cell loss), and the overall response was comparable in all laboratories despite some differences in doses tested. The results of the collaborative study for the coded compounds were generally reproducible among the laboratories involved and intra-laboratory reproducibility was also good. These data indicate that the comet assay in EpiDerm™ skin models is a promising model for the safety assessment of compounds with a dermal route of exposure.

Inter-laboratory quality control for hormone-dependent gene expression in human breast tumors using real-time reverse transcription-polymerase chain reaction.

PubMed

de Cremoux, P; Bieche, I; Tran-Perennou, C; Vignaud, S; Boudou, E; Asselain, B; Lidereau, R; Magdelénat, H; Becette, V; Sigal-Zafrani, B; Spyratos, F

2004-09-01

Quantitative reverse transcription-polymerase chain reaction (RT-PCR) used to detect minor changes in specific mRNA concentrations may be associated with poor reproducibility. Stringent quality control is therefore essential at each step of the protocol, including the PCR procedure. We performed inter-laboratory quality control of quantitative PCR between two independent laboratories, using in-house RT-PCR assays on a series of hormone-related target genes in a retrospective consecutive series of 79 breast tumors. Total RNA was reverse transcribed in a single center. Calibration curves were performed for five target genes (estrogen receptor (ER)alpha, ERbeta, progesterone receptor (PR), CYP19 (aromatase) and Ki 67) and for two reference genes (human acidic ribosomal phosphoprotein PO (RPLPO) and TATA box-binding protein (TBP)). Amplification efficiencies of the calibrator were determined for each run and used to calculate mRNA expression. Correlation coefficients were evaluated for each target and each reference gene. A good correlation was observed for all target and reference genes in both centers using their own protocols and kits (P < 0.0001). The correlation coefficients ranged from 0.90 to 0.98 for the various target genes in the two centers. A good correlation was observed between the level of expression of the ERalpha and the PR transcripts (P < 0.001). A weak inverse correlation was observed in both centers between ERalpha and ERbeta levels, but only when TBP was the reference gene. No other correlation was observed with other parameters. Real-time PCR assays allow convenient quantification of target mRNA transcripts and quantification of target-derived nucleic acids in clinical specimens. This study addresses the importance of inter-laboratory quality controls for the use of a panel of real-time PCR assays devoted to clinical samples and protocols and to ensure their appropriate accuracy. This can also facilitate exchanges and multicenter comparison of data.

Comet assay in reconstructed 3D human epidermal skin models—investigation of intra- and inter-laboratory reproducibility with coded chemicals

PubMed Central

Pfuhler, Stefan

2013-01-01

Reconstructed 3D human epidermal skin models are being used increasingly for safety testing of chemicals. Based on EpiDerm™ tissues, an assay was developed in which the tissues were topically exposed to test chemicals for 3h followed by cell isolation and assessment of DNA damage using the comet assay. Inter-laboratory reproducibility of the 3D skin comet assay was initially demonstrated using two model genotoxic carcinogens, methyl methane sulfonate (MMS) and 4-nitroquinoline-n-oxide, and the results showed good concordance among three different laboratories and with in vivo data. In Phase 2 of the project, intra- and inter-laboratory reproducibility was investigated with five coded compounds with different genotoxicity liability tested at three different laboratories. For the genotoxic carcinogens MMS and N-ethyl-N-nitrosourea, all laboratories reported a dose-related and statistically significant increase (P < 0.05) in DNA damage in every experiment. For the genotoxic carcinogen, 2,4-diaminotoluene, the overall result from all laboratories showed a smaller, but significant genotoxic response (P < 0.05). For cyclohexanone (CHN) (non-genotoxic in vitro and in vivo, and non-carcinogenic), an increase compared to the solvent control acetone was observed only in one laboratory. However, the response was not dose related and CHN was judged negative overall, as was p-nitrophenol (p-NP) (genotoxic in vitro but not in vivo and non-carcinogenic), which was the only compound showing clear cytotoxic effects. For p-NP, significant DNA damage generally occurred only at doses that were substantially cytotoxic (>30% cell loss), and the overall response was comparable in all laboratories despite some differences in doses tested. The results of the collaborative study for the coded compounds were generally reproducible among the laboratories involved and intra-laboratory reproducibility was also good. These data indicate that the comet assay in EpiDerm™ skin models is a promising model for the safety assessment of compounds with a dermal route of exposure. PMID:24150594

Progress in the application of DNA microarrays.

PubMed Central

Lobenhofer, E K; Bushel, P R; Afshari, C A; Hamadeh, H K

2001-01-01

Microarray technology has been applied to a variety of different fields to address fundamental research questions. The use of microarrays, or DNA chips, to study the gene expression profiles of biologic samples began in 1995. Since that time, the fundamental concepts behind the chip, the technology required for making and using these chips, and the multitude of statistical tools for analyzing the data have been extensively reviewed. For this reason, the focus of this review will be not on the technology itself but on the application of microarrays as a research tool and the future challenges of the field. PMID:11673116

Constructing Tissue Microarrays: Protocols and Methods Considering Potential Advantages and Disadvantages for Downstream Use.

PubMed

Bingle, Lynne; Fonseca, Felipe P; Farthing, Paula M

2017-01-01

Tissue microarrays were first constructed in the 1980s but were used by only a limited number of researchers for a considerable period of time. In the last 10 years there has been a dramatic increase in the number of publications describing the successful use of tissue microarrays in studies aimed at discovering and validating biomarkers. This, along with the increased availability of both manual and automated microarray builders on the market, has encouraged even greater use of this novel and powerful tool. This chapter describes the basic techniques required to build a tissue microarray using a manual method in order that the theory behind the practical steps can be fully explained. Guidance is given to ensure potential disadvantages of the technique are fully considered.

Interlaboratory study of a method for determining nonvolatile organic carbon in aquifer materials

USGS Publications Warehouse

Caughey, M.E.; Barcelona, M.J.; Powell, R.M.; Cahill, R.A.; Gron, C.; Lawrenz, D.; Meschi, P.L.

1995-01-01

The organic carbon fraction in aquifer materials exerts a major influence on the subsurface mobilities of organic and organic-associated contaminants. The spatial distribution of total organic carbon (TOC) in aquifer materials must be determined before the transport of hydrophobic organic pollutants in aquifers can be modeled accurately. Previous interlaboratory studies showed that it is difficult to measure TOC concentrations 1%. We have tested a new analytical method designed to improve the accuracy and precision of nonvolatile TOC quantitation in geologic materials that also contain carbonate minerals. Four authentic aquifer materials and one NIST standard reference material were selected as test materials for a blind collaborative study. Nonvolatile TOC in these materials ranged from 0.05 to 1.4%, while TIC ranged from 0.46 to 12.6%. Sample replicates were digested with sulfurous acid, dried at 40??C, and then combusted at 950??C using LECO or UIC instruments. For the three test materials that contained >2% TIC, incomplete acidification resulted in a systematic positive bias of TOC values reported by five of the six laboratories that used the test method. Participants did not have enough time to become proficient with the new method before they analyzed the test materials. A seventh laboratory successfully used an alternative method that analyzed separate liquid and solid fractions of the acidified sample residues. ?? 1995 Springer-Verlag.

Round-robin study of arsenic implant dose measurement in silicon by SIMS

NASA Astrophysics Data System (ADS)

Simons, D.; Kim, K.; Benbalagh, R.; Bennett, J.; Chew, A.; Gehre, D.; Hasegawa, T.; Hitzman, C.; Ko, J.; Lindstrom, R.; MacDonald, B.; Magee, C.; Montgomery, N.; Peres, P.; Ronsheim, P.; Yoshikawa, S.; Schuhmacher, M.; Stockwell, W.; Sykes, D.; Tomita, M.; Toujou, F.; Won, J.

2006-07-01

An international round-robin study was undertaken under the auspices of ISO TC201/SC6 to determine the best analytical conditions and the level of interlaboratory agreement for the determination of the implantation dose of arsenic in silicon by secondary ion mass spectrometry (SIMS). Fifteen SIMS laboratories, as well as two laboratories that performed low energy electron-induced X-ray emission spectrometry (LEXES) and one that made measurements by instrumental neutron activation analysis (INAA) were asked to determine the implanted arsenic doses in three unknown samples using as a comparator NIST Standard Reference Material ® 2134. The use of a common reference material by all laboratories resulted in better interlaboratory agreement than was seen in a previous round-robin that lacked a common comparator. The relative standard deviation among laboratories was less than 4% for the medium-dose sample, but several percent larger for the low- and high-dose samples. The high-dose sample showed a significant difference between point-by-point and average matrix normalization because the matrix signal decreased in the vicinity of the implant peak, as observed in a previous study. The dose from point-by-point normalization was in close agreement with that determined by INAA. No clear difference in measurement repeatability was seen when comparing Si 2- and Si 3- as matrix references with AsSi -.

Examination of a size-change test for photovoltaic encapsulation materials

NASA Astrophysics Data System (ADS)

Miller, David C.; Gu, Xiaohong; Ji, Liang; Kelly, George; Nickel, Nichole; Norum, Paul; Shioda, Tsuyoshi; Tamizhmani, Govindasamy; Wohlgemuth, John H.

2012-10-01

We examine a proposed test standard that can be used to evaluate the maximum representative change in linear dimensions of sheet encapsulation products for photovoltaic modules (resulting from their thermal processing). The proposed protocol is part of a series of material-level tests being developed within Working Group 2 of the Technical Committee 82 of the International Electrotechnical Commission. The characterization tests are being developed to aid module design (by identifying the essential characteristics that should be communicated on a datasheet), quality control (via internal material acceptance and process control), and failure analysis. Discovery and interlaboratory experiments were used to select particular parameters for the size-change test. The choice of a sand substrate and aluminum carrier is explored relative to other options. The temperature uniformity of +/-5°C for the substrate was confirmed using thermography. Considerations related to the heating device (hot-plate or oven) are explored. The time duration of 5 minutes was identified from the time-series photographic characterization of material specimens (EVA, ionomer, PVB, TPO, and TPU). The test procedure was revised to account for observed effects of size and edges. The interlaboratory study identified typical size-change characteristics, and also verified the absolute reproducibility of +/-5% between laboratories.

Canine and feline hematology reference values for the ADVIA 120 hematology system.

PubMed

Moritz, Andreas; Fickenscher, Yvonne; Meyer, Karin; Failing, Klaus; Weiss, Douglas J

2004-01-01

The ADVIA 120 is a laser-based hematology analyzer with software applications for animal species. Accurate reference values would be useful for the assessment of new hematologic parameters and for interlaboratory comparisons. The goal of this study was to establish reference intervals for CBC results and new parameters for RBC morphology, reticulocytes, and platelets in healthy dogs and cats using the ADVIA 120 hematology system. The ADVIA 120, with multispecies software (version 1.107-MS), was used to analyze whole blood samples from clinically healthy dogs (n=46) and cats (n=61). Data distribution was determined and reference intervals were calculated as 2.5 to 97.5 percentiles and 25 to 75 percentiles. Most data showed Gaussian or log-normal distribution. The numbers of RBCs falling outside the normocytic-normochromic range were slightly higher in cats than in dogs. Both dogs and cats had reticulocytes with low, medium, and high absorbance. Mean numbers of large platelets and platelet clumps were higher in cats compared with dogs. Reference intervals obtained on the ADVIA 120 provide valuable baseline information for assessing new hematologic parameters and for interlaboratory comparisons. Differences compared with previously published reference values can be attributed largely to differences in methodology.

Comparison of δ18O measurements in nitrate by different combustion techniques

USGS Publications Warehouse

Revesz, Kinga; Böhlke, John Karl

2002-01-01

Three different KNO3 salts with δ18O values ranging from about −31 to +54‰ relative to VSMOW were used to compare three off-line, sealed glass tube combustion methods (widely used for isotope studies) with a more recently developed on-line carbon combustion technique. All methods yielded roughly similar isotope ratios for KNO3 samples with δ18O values in the midpoint of the δ18O scale near that of the nitrate reference material IAEA-NO-3 (around +21 to +25‰). This reference material has been used previously for one-point interlaboratory and intertechnique calibrations. However, the isotope ratio scale factors by all of the off-line combustion techniques are compressed such that they are between 0.3 and 0.7 times that of the on-line combustion technique. The contraction of the δ18O scale in the off-line preparations apparently is caused by O isotope exchange between the sample and the glass combustion tubes. These results reinforce the need for nitrate reference materials with δ18O values far from that of atmospheric O2, to improve interlaboratory comparability.

CEM-designer: design of custom expression microarrays in the post-ENCODE Era.

PubMed

Arnold, Christian; Externbrink, Fabian; Hackermüller, Jörg; Reiche, Kristin

2014-11-10

Microarrays are widely used in gene expression studies, and custom expression microarrays are popular to monitor expression changes of a customer-defined set of genes. However, the complexity of transcriptomes uncovered recently make custom expression microarray design a non-trivial task. Pervasive transcription and alternative processing of transcripts generate a wealth of interweaved transcripts that requires well-considered probe design strategies and is largely neglected in existing approaches. We developed the web server CEM-Designer that facilitates microarray platform independent design of custom expression microarrays for complex transcriptomes. CEM-Designer covers (i) the collection and generation of a set of unique target sequences from different sources and (ii) the selection of a set of sensitive and specific probes that optimally represents the target sequences. Probe design itself is left to third party software to ensure that probes meet provider-specific constraints. CEM-Designer is available at http://designpipeline.bioinf.uni-leipzig.de. Copyright © 2014 Elsevier B.V. All rights reserved.

The effect of column purification on cDNA indirect labelling for microarrays

PubMed Central

Molas, M Lia; Kiss, John Z

2007-01-01

Background The success of the microarray reproducibility is dependent upon the performance of standardized procedures. Since the introduction of microarray technology for the analysis of global gene expression, reproducibility of results among different laboratories has been a major problem. Two of the main contributors to this variability are the use of different microarray platforms and different laboratory practices. In this paper, we address the latter question in terms of how variation in one of the steps of a labelling procedure affects the cDNA product prior to microarray hybridization. Results We used a standard procedure to label cDNA for microarray hybridization and employed different types of column chromatography for cDNA purification. After purifying labelled cDNA, we used the Agilent 2100 Bioanalyzer and agarose gel electrophoresis to assess the quality of the labelled cDNA before its hybridization onto a microarray platform. There were major differences in the cDNA profile (i.e. cDNA fragment lengths and abundance) as a result of using four different columns for purification. In addition, different columns have different efficiencies to remove rRNA contamination. This study indicates that the appropriate column to use in this type of protocol has to be experimentally determined. Finally, we present new evidence establishing the importance of testing the method of purification used during an indirect labelling procedure. Our results confirm the importance of assessing the quality of the sample in the labelling procedure prior to hybridization onto a microarray platform. Conclusion Standardization of column purification systems to be used in labelling procedures will improve the reproducibility of microarray results among different laboratories. In addition, implementation of a quality control check point of the labelled samples prior to microarray hybridization will prevent hybridizing a poor quality sample to expensive micorarrays. PMID:17597522

The effect of column purification on cDNA indirect labelling for microarrays.

PubMed

Molas, M Lia; Kiss, John Z

2007-06-27

The success of the microarray reproducibility is dependent upon the performance of standardized procedures. Since the introduction of microarray technology for the analysis of global gene expression, reproducibility of results among different laboratories has been a major problem. Two of the main contributors to this variability are the use of different microarray platforms and different laboratory practices. In this paper, we address the latter question in terms of how variation in one of the steps of a labelling procedure affects the cDNA product prior to microarray hybridization. We used a standard procedure to label cDNA for microarray hybridization and employed different types of column chromatography for cDNA purification. After purifying labelled cDNA, we used the Agilent 2100 Bioanalyzer and agarose gel electrophoresis to assess the quality of the labelled cDNA before its hybridization onto a microarray platform. There were major differences in the cDNA profile (i.e. cDNA fragment lengths and abundance) as a result of using four different columns for purification. In addition, different columns have different efficiencies to remove rRNA contamination. This study indicates that the appropriate column to use in this type of protocol has to be experimentally determined. Finally, we present new evidence establishing the importance of testing the method of purification used during an indirect labelling procedure. Our results confirm the importance of assessing the quality of the sample in the labelling procedure prior to hybridization onto a microarray platform. Standardization of column purification systems to be used in labelling procedures will improve the reproducibility of microarray results among different laboratories. In addition, implementation of a quality control check point of the labelled samples prior to microarray hybridization will prevent hybridizing a poor quality sample to expensive micorarrays.

Evolution of the MIDTAL microarray: the adaption and testing of oligonucleotide 18S and 28S rDNA probes and evaluation of subsequent microarray generations with Prymnesium spp. cultures and field samples.

PubMed

McCoy, Gary R; Touzet, Nicolas; Fleming, Gerard T A; Raine, Robin

2015-07-01

The toxic microalgal species Prymnesium parvum and Prymnesium polylepis are responsible for numerous fish kills causing economic stress on the aquaculture industry and, through the consumption of contaminated shellfish, can potentially impact on human health. Monitoring of toxic phytoplankton is traditionally carried out by light microscopy. However, molecular methods of identification and quantification are becoming more common place. This study documents the optimisation of the novel Microarrays for the Detection of Toxic Algae (MIDTAL) microarray from its initial stages to the final commercial version now available from Microbia Environnement (France). Existing oligonucleotide probes used in whole-cell fluorescent in situ hybridisation (FISH) for Prymnesium species from higher group probes to species-level probes were adapted and tested on the first-generation microarray. The combination and interaction of numerous other probes specific for a whole range of phytoplankton taxa also spotted on the chip surface caused high cross reactivity, resulting in false-positive results on the microarray. The probe sequences were extended for the subsequent second-generation microarray, and further adaptations of the hybridisation protocol and incubation temperatures significantly reduced false-positive readings from the first to the second-generation chip, thereby increasing the specificity of the MIDTAL microarray. Additional refinement of the subsequent third-generation microarray protocols with the addition of a poly-T amino linker to the 5' end of each probe further enhanced the microarray performance but also highlighted the importance of optimising RNA labelling efficiency when testing with natural seawater samples from Killary Harbour, Ireland.

Characterization and simulation of cDNA microarray spots using a novel mathematical model

PubMed Central

Kim, Hye Young; Lee, Seo Eun; Kim, Min Jung; Han, Jin Il; Kim, Bo Kyung; Lee, Yong Sung; Lee, Young Seek; Kim, Jin Hyuk

2007-01-01

Background The quality of cDNA microarray data is crucial for expanding its application to other research areas, such as the study of gene regulatory networks. Despite the fact that a number of algorithms have been suggested to increase the accuracy of microarray gene expression data, it is necessary to obtain reliable microarray images by improving wet-lab experiments. As the first step of a cDNA microarray experiment, spotting cDNA probes is critical to determining the quality of spot images. Results We developed a governing equation of cDNA deposition during evaporation of a drop in the microarray spotting process. The governing equation included four parameters: the surface site density on the support, the extrapolated equilibrium constant for the binding of cDNA molecules with surface sites on glass slides, the macromolecular interaction factor, and the volume constant of a drop of cDNA solution. We simulated cDNA deposition from the single model equation by varying the value of the parameters. The morphology of the resulting cDNA deposit can be classified into three types: a doughnut shape, a peak shape, and a volcano shape. The spot morphology can be changed into a flat shape by varying the experimental conditions while considering the parameters of the governing equation of cDNA deposition. The four parameters were estimated by fitting the governing equation to the real microarray images. With the results of the simulation and the parameter estimation, the phenomenon of the formation of cDNA deposits in each type was investigated. Conclusion This study explains how various spot shapes can exist and suggests which parameters are to be adjusted for obtaining a good spot. This system is able to explore the cDNA microarray spotting process in a predictable, manageable and descriptive manner. We hope it can provide a way to predict the incidents that can occur during a real cDNA microarray experiment, and produce useful data for several research applications involving cDNA microarrays. PMID:18096047

Single-Nucleotide Polymorphism-Microarray Ploidy Analysis of Paraffin-Embedded Products of Conception in Recurrent Pregnancy Loss Evaluations.

PubMed

Maslow, Bat-Sheva L; Budinetz, Tara; Sueldo, Carolina; Anspach, Erica; Engmann, Lawrence; Benadiva, Claudio; Nulsen, John C

2015-07-01

To compare the analysis of chromosome number from paraffin-embedded products of conception using single-nucleotide polymorphism (SNP) microarray with the recommended screening for the evaluation of couples presenting with recurrent pregnancy loss who do not have previous fetal cytogenetic data. We performed a retrospective cohort study including all women who presented for a new evaluation of recurrent pregnancy loss over a 2-year period (January 1, 2012, to December 31, 2013). All participants had at least two documented first-trimester losses and both the recommended screening tests and SNP microarray performed on at least one paraffin-embedded products of conception sample. Single-nucleotide polymorphism microarray identifies all 24 chromosomes (22 autosomes, X, and Y). Forty-two women with a total of 178 losses were included in the study. Paraffin-embedded products of conception from 62 losses were sent for SNP microarray. Single-nucleotide polymorphism microarray successfully diagnosed fetal chromosome number in 71% (44/62) of samples, of which 43% (19/44) were euploid and 57% (25/44) were noneuploid. Seven of 42 (17%) participants had abnormalities on recurrent pregnancy loss screening. The per-person detection rate for a cause of pregnancy loss was significantly higher in the SNP microarray (0.50; 95% confidence interval [CI] 0.36-0.64) compared with recurrent pregnancy loss evaluation (0.17; 95% CI 0.08-0.31) (P=.002). Participants with one or more euploid loss identified on paraffin-embedded products of conception were significantly more likely to have an abnormality on recurrent pregnancy loss screening than those with only noneuploid results (P=.028). The significance remained when controlling for age, number of losses, number of samples, and total pregnancies. These results suggest that SNP microarray testing of paraffin-embedded products of conception is a valuable tool for the evaluation of recurrent pregnancy loss in patients without prior fetal cytogenetic results. Recommended recurrent pregnancy loss screening was unnecessary in almost half the patients in our study. II.

The Choice of the Filtering Method in Microarrays Affects the Inference Regarding Dosage Compensation of the Active X-Chromosome

PubMed Central

Zeller, Tanja; Wild, Philipp S.; Truong, Vinh; Trégouët, David-Alexandre; Munzel, Thomas; Ziegler, Andreas; Cambien, François; Blankenberg, Stefan; Tiret, Laurence

2011-01-01

Background The hypothesis of dosage compensation of genes of the X chromosome, supported by previous microarray studies, was recently challenged by RNA-sequencing data. It was suggested that microarray studies were biased toward an over-estimation of X-linked expression levels as a consequence of the filtering of genes below the detection threshold of microarrays. Methodology/Principal Findings To investigate this hypothesis, we used microarray expression data from circulating monocytes in 1,467 individuals. In total, 25,349 and 1,156 probes were unambiguously assigned to autosomes and the X chromosome, respectively. Globally, there was a clear shift of X-linked expressions toward lower levels than autosomes. We compared the ratio of expression levels of X-linked to autosomal transcripts (X∶AA) using two different filtering methods: 1. gene expressions were filtered out using a detection threshold irrespective of gene chromosomal location (the standard method in microarrays); 2. equal proportions of genes were filtered out separately on the X and on autosomes. For a wide range of filtering proportions, the X∶AA ratio estimated with the first method was not significantly different from 1, the value expected if dosage compensation was achieved, whereas it was significantly lower than 1 with the second method, leading to the rejection of the hypothesis of dosage compensation. We further showed in simulated data that the choice of the most appropriate method was dependent on biological assumptions regarding the proportion of actively expressed genes on the X chromosome comparative to the autosomes and the extent of dosage compensation. Conclusion/Significance This study shows that the method used for filtering out lowly expressed genes in microarrays may have a major impact according to the hypothesis investigated. The hypothesis of dosage compensation of X-linked genes cannot be firmly accepted or rejected using microarray-based data. PMID:21912656

A genome-wide 20 K citrus microarray for gene expression analysis

PubMed Central

Martinez-Godoy, M Angeles; Mauri, Nuria; Juarez, Jose; Marques, M Carmen; Santiago, Julia; Forment, Javier; Gadea, Jose

2008-01-01

Background Understanding of genetic elements that contribute to key aspects of citrus biology will impact future improvements in this economically important crop. Global gene expression analysis demands microarray platforms with a high genome coverage. In the last years, genome-wide EST collections have been generated in citrus, opening the possibility to create new tools for functional genomics in this crop plant. Results We have designed and constructed a publicly available genome-wide cDNA microarray that include 21,081 putative unigenes of citrus. As a functional companion to the microarray, a web-browsable database [1] was created and populated with information about the unigenes represented in the microarray, including cDNA libraries, isolated clones, raw and processed nucleotide and protein sequences, and results of all the structural and functional annotation of the unigenes, like general description, BLAST hits, putative Arabidopsis orthologs, microsatellites, putative SNPs, GO classification and PFAM domains. We have performed a Gene Ontology comparison with the full set of Arabidopsis proteins to estimate the genome coverage of the microarray. We have also performed microarray hybridizations to check its usability. Conclusion This new cDNA microarray replaces the first 7K microarray generated two years ago and allows gene expression analysis at a more global scale. We have followed a rational design to minimize cross-hybridization while maintaining its utility for different citrus species. Furthermore, we also provide access to a website with full structural and functional annotation of the unigenes represented in the microarray, along with the ability to use this site to directly perform gene expression analysis using standard tools at different publicly available servers. Furthermore, we show how this microarray offers a good representation of the citrus genome and present the usefulness of this genomic tool for global studies in citrus by using it to catalogue genes expressed in citrus globular embryos. PMID:18598343

A study of metaheuristic algorithms for high dimensional feature selection on microarray data

NASA Astrophysics Data System (ADS)

Dankolo, Muhammad Nasiru; Radzi, Nor Haizan Mohamed; Sallehuddin, Roselina; Mustaffa, Noorfa Haszlinna

2017-11-01

Microarray systems enable experts to examine gene profile at molecular level using machine learning algorithms. It increases the potentials of classification and diagnosis of many diseases at gene expression level. Though, numerous difficulties may affect the efficiency of machine learning algorithms which includes vast number of genes features comprised in the original data. Many of these features may be unrelated to the intended analysis. Therefore, feature selection is necessary to be performed in the data pre-processing. Many feature selection algorithms are developed and applied on microarray which including the metaheuristic optimization algorithms. This paper discusses the application of the metaheuristics algorithms for feature selection in microarray dataset. This study reveals that, the algorithms have yield an interesting result with limited resources thereby saving computational expenses of machine learning algorithms.

The use of open source bioinformatics tools to dissect transcriptomic data.

PubMed

Nitsche, Benjamin M; Ram, Arthur F J; Meyer, Vera

2012-01-01

Microarrays are a valuable technology to study fungal physiology on a transcriptomic level. Various microarray platforms are available comprising both single and two channel arrays. Despite different technologies, preprocessing of microarray data generally includes quality control, background correction, normalization, and summarization of probe level data. Subsequently, depending on the experimental design, diverse statistical analysis can be performed, including the identification of differentially expressed genes and the construction of gene coexpression networks.We describe how Bioconductor, a collection of open source and open development packages for the statistical programming language R, can be used for dissecting microarray data. We provide fundamental details that facilitate the process of getting started with R and Bioconductor. Using two publicly available microarray datasets from Aspergillus niger, we give detailed protocols on how to identify differentially expressed genes and how to construct gene coexpression networks.

CYANOBACTERIA AND CYANOTOXINS IN WATER SUPPLY RESERVOIRS – TO DEVELOP AND VALIDATE A MICROARRAY TO TEST FOR CYANOBACTERIA AND CYANOTOXIN GENES IN DRINKING WATER RESERVOIRS AS AN AID TO RISK ASSESSMENT AND MANAGEMENT OF WATER SUPPLIES

EPA Science Inventory

The objective of this study is to develop a microarray to test for cyanobacteria and cyanotoxin genes in drinking water reservoirs as an aid to risk assessment and manages of water supplies. The microarray will include probes recognizing important freshwater cyanobacterial tax...

Microarray platform affords improved product analysis in mammalian cell growth studies

PubMed Central

Li, Lingyun; Migliore, Nicole; Schaefer, Eugene; Sharfstein, Susan T.; Dordick, Jonathan S.; Linhardt, Robert J.

2014-01-01

High throughput (HT) platforms serve as cost-efficient and rapid screening method for evaluating the effect of cell culture conditions and screening of chemicals. The aim of the current study was to develop a high-throughput cell-based microarray platform to assess the effect of culture conditions on Chinese hamster ovary (CHO) cells. Specifically, growth, transgene expression and metabolism of a GS/MSX CHO cell line, which produces a therapeutic monoclonal antibody, was examined using microarray system in conjunction with conventional shake flask platform in a non-proprietary medium. The microarray system consists of 60 nl spots of cells encapsulated in alginate and separated in groups via an 8-well chamber system attached to the chip. Results show the non-proprietary medium developed allows cell growth, production and normal glycosylation of recombinant antibody and metabolism of the recombinant CHO cells in both the microarray and shake flask platforms. In addition, 10.3 mM glutamate addition to the defined base media results in lactate metabolism shift in the recombinant GS/MSX CHO cells in the shake flask platform. Ultimately, the results demonstrate that the high-throughput microarray platform has the potential to be utilized for evaluating the impact of media additives on cellular processes, such as, cell growth, metabolism and productivity. PMID:24227746

Emerging Use of Gene Expression Microarrays in Plant Physiology

DOE PAGES

Wullschleger, Stan D.; Difazio, Stephen P.

2003-01-01

Microarrays have become an important technology for the global analysis of gene expression in humans, animals, plants, and microbes. Implemented in the context of a well-designed experiment, cDNA and oligonucleotide arrays can provide highthroughput, simultaneous analysis of transcript abundance for hundreds, if not thousands, of genes. However, despite widespread acceptance, the use of microarrays as a tool to better understand processes of interest to the plant physiologist is still being explored. To help illustrate current uses of microarrays in the plant sciences, several case studies that we believe demonstrate the emerging application of gene expression arrays in plant physiology weremore » selected from among the many posters and presentations at the 2003 Plant and Animal Genome XI Conference. Based on this survey, microarrays are being used to assess gene expression in plants exposed to the experimental manipulation of air temperature, soil water content and aluminium concentration in the root zone. Analysis often includes characterizing transcript profiles for multiple post-treatment sampling periods and categorizing genes with common patterns of response using hierarchical clustering techniques. In addition, microarrays are also providing insights into developmental changes in gene expression associated with fibre and root elongation in cotton and maize, respectively. Technical and analytical limitations of microarrays are discussed and projects attempting to advance areas of microarray design and data analysis are highlighted. Finally, although much work remains, we conclude that microarrays are a valuable tool for the plant physiologist interested in the characterization and identification of individual genes and gene families with potential application in the fields of agriculture, horticulture and forestry.« less

Profiling In Situ Microbial Community Structure with an Amplification Microarray

PubMed Central

Knickerbocker, Christopher; Bryant, Lexi; Golova, Julia; Wiles, Cory; Williams, Kenneth H.; Peacock, Aaron D.; Long, Philip E.

2013-01-01

The objectives of this study were to unify amplification, labeling, and microarray hybridization chemistries within a single, closed microfluidic chamber (an amplification microarray) and verify technology performance on a series of groundwater samples from an in situ field experiment designed to compare U(VI) mobility under conditions of various alkalinities (as HCO3−) during stimulated microbial activity accompanying acetate amendment. Analytical limits of detection were between 2 and 200 cell equivalents of purified DNA. Amplification microarray signatures were well correlated with 16S rRNA-targeted quantitative PCR results and hybridization microarray signatures. The succession of the microbial community was evident with and consistent between the two microarray platforms. Amplification microarray analysis of acetate-treated groundwater showed elevated levels of iron-reducing bacteria (Flexibacter, Geobacter, Rhodoferax, and Shewanella) relative to the average background profile, as expected. Identical molecular signatures were evident in the transect treated with acetate plus NaHCO3, but at much lower signal intensities and with a much more rapid decline (to nondetection). Azoarcus, Thaurea, and Methylobacterium were responsive in the acetate-only transect but not in the presence of bicarbonate. Observed differences in microbial community composition or response to bicarbonate amendment likely had an effect on measured rates of U reduction, with higher rates probable in the part of the field experiment that was amended with bicarbonate. The simplification in microarray-based work flow is a significant technological advance toward entirely closed-amplicon microarray-based tests and is generally extensible to any number of environmental monitoring applications. PMID:23160129

Deciphering the glycosaminoglycan code with the help of microarrays.

PubMed

de Paz, Jose L; Seeberger, Peter H

2008-07-01

Carbohydrate microarrays have become a powerful tool to elucidate the biological role of complex sugars. Microarrays are particularly useful for the study of glycosaminoglycans (GAGs), a key class of carbohydrates. The high-throughput chip format enables rapid screening of large numbers of potential GAG sequences produced via a complex biosynthesis while consuming very little sample. Here, we briefly highlight the most recent advances involving GAG microarrays built with synthetic or naturally derived oligosaccharides. These chips are powerful tools for characterizing GAG-protein interactions and determining structure-activity relationships for specific sequences. Thereby, they contribute to decoding the information contained in specific GAG sequences.

Fibre optic microarrays.

PubMed

Walt, David R

2010-01-01

This tutorial review describes how fibre optic microarrays can be used to create a variety of sensing and measurement systems. This review covers the basics of optical fibres and arrays, the different microarray architectures, and describes a multitude of applications. Such arrays enable multiplexed sensing for a variety of analytes including nucleic acids, vapours, and biomolecules. Polymer-coated fibre arrays can be used for measuring microscopic chemical phenomena, such as corrosion and localized release of biochemicals from cells. In addition, these microarrays can serve as a substrate for fundamental studies of single molecules and single cells. The review covers topics of interest to chemists, biologists, materials scientists, and engineers.

Evaluation of cross-connected waveguides as transfer standards of transmission at high millimetre-wave frequencies

NASA Astrophysics Data System (ADS)

Ridler, Nick; Clarke, Roland; Huang, Hui; Zinal, Sherko

2016-08-01

At the present time, transfer and verification standards of transmission coefficient (or, equivalently, transmission loss) are not readily available at high millimetre-wave frequencies (i.e. at frequencies ranging typically from 100 GHz to 300 GHz). In recent years, cross-connected waveguide devices have been proposed to provide calculable standards of transmission loss at these frequencies. This paper investigates the viability of these cross-connected waveguides as transfer standards of transmission for inter-laboratory measurement comparison exercises. This relates to their potential use in activities such as international key comparison exercises and measurement audit programmes. A trial inter-laboratory comparison involving four laboratories using two cross-connected waveguides in the WR-05 waveguide size (covering frequencies from 140 GHz to 220 GHz) is described and includes an analysis of the measurement results obtained during the comparison exercise.

Inter-laboratory comparisons of short-lived gamma-emitting radionuclides in nuclear reactor water.

PubMed

Klemola, S K

2008-01-01

Inter-laboratory comparisons of gamma-emitting nuclides in nuclear power plant coolant water have been carried out in Finland since 1994. The reactor water samples are taken and prepared by one of the two nuclear power plants and delivered to the participants. Since all the participants get their sample within just a few hours it has been possible to analyse and compare results of nuclides with half-lives shorter than 1h. The total number of short-lived nuclides is 26. All the main nuclides are regularly identified and the activities have been obtained with reasonable accuracy throughout the years. The overall deviation of the results has decreased in 13 years. The effects of true coincidence summing and discrepancies in nuclear data have been identified as potential sources of remaining discrepancies. All the participants have found this type of comparison very useful.

(60)Co in cast steel matrix: A European interlaboratory comparison for the characterisation of new activity standards for calibration of gamma-ray spectrometers in metallurgy.

PubMed

Tzika, Faidra; Burda, Oleksiy; Hult, Mikael; Arnold, Dirk; Marroyo, Belén Caro; Dryák, Pavel; Fazio, Aldo; Ferreux, Laurent; García-Toraño, Eduardo; Javornik, Andrej; Klemola, Seppo; Luca, Aurelian; Moser, Hannah; Nečemer, Marijan; Peyrés, Virginia; Reis, Mario; Silva, Lidia; Šolc, Jaroslav; Svec, Anton; Tyminski, Zbigniew; Vodenik, Branko; Wätjen, Uwe

2016-08-01

Two series of activity standards of (60)Co in cast steel matrix, developed for the calibration of gamma-ray spectrometry systems in the metallurgical sector, were characterised using a European interlaboratory comparison among twelve National Metrology Institutes and one international organisation. The first standard, consisting of 14 disc shaped samples, was cast from steel contaminated during production ("originally"), and the second, consisting of 15 similar discs, from artificially-contaminated ("spiked") steel. The reference activity concentrations of (60)Co in the cast steel standards were (1.077±0.019) Bqg(-1) on 1 January 2013 12h00 UT and (1.483±0.022) Bqg(-1) on 1 June 2013 12h00 UT, respectively. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Quality assurance in RT-PCR-based BCR/ABL diagnostics--results of an interlaboratory test and a standardization approach.

PubMed

Burmeister, T; Maurer, J; Aivado, M; Elmaagacli, A H; Grünebach, F; Held, K R; Hess, G; Hochhaus, A; Höppner, W; Lentes, K U; Lübbert, M; Schäfer, K L; Schafhausen, P; Schmidt, C A; Schüler, F; Seeger, K; Seelig, R; Thiede, C; Viehmann, S; Weber, C; Wilhelm, S; Christmann, A; Clement, J H; Ebener, U; Enczmann, J; Leo, R; Schleuning, M; Schoch, R; Thiel, E

2000-10-01

Here we describe the results of an interlaboratory test for RT-PCR-based BCR/ABL analysis. The test was organized in two parts. The number of participating laboratories in the first and second part was 27 and 20, respectively. In the first part samples containing various concentrations of plasmids with the ela2, b2a2 or b3a2 BCR/ABL transcripts were analyzed by PCR. In the second part of the test, cell samples containing various concentrations of BCR/ABL-positive cells were analyzed by RT-PCR. Overall PCR sensitivity was sufficient in approximately 90% of the tests, but a significant number of false positive results were obtained. There were significant differences in sensitivity in the cell-based analysis between the various participants. The results are discussed, and proposals are made regarding the choice of primers, controls, conditions for RNA extraction and reverse transcription.

Transfer Standard Uncertainty Can Cause Inconclusive Inter-Laboratory Comparisons

PubMed Central

Wright, John; Toman, Blaza; Mickan, Bodo; Wübbeler, Gerd; Bodnar, Olha; Elster, Clemens

2016-01-01

Inter-laboratory comparisons use the best available transfer standards to check the participants’ uncertainty analyses, identify underestimated uncertainty claims or unknown measurement biases, and improve the global measurement system. For some measurands, instability of the transfer standard can lead to an inconclusive comparison result. If the transfer standard uncertainty is large relative to a participating laboratory’s uncertainty, the commonly used standardized degree of equivalence ≤ 1 criterion does not always correctly assess whether a participant is working within their uncertainty claims. We show comparison results that demonstrate this issue and propose several criteria for assessing a comparison result as passing, failing, or inconclusive. We investigate the behavior of the standardized degree of equivalence and alternative comparison measures for a range of values of the transfer standard uncertainty relative to the individual laboratory uncertainty values. The proposed alternative criteria successfully discerned between passing, failing, and inconclusive comparison results for the cases we examined. PMID:28090123

Automatic Identification and Quantification of Extra-Well Fluorescence in Microarray Images.

PubMed

Rivera, Robert; Wang, Jie; Yu, Xiaobo; Demirkan, Gokhan; Hopper, Marika; Bian, Xiaofang; Tahsin, Tasnia; Magee, D Mitchell; Qiu, Ji; LaBaer, Joshua; Wallstrom, Garrick

2017-11-03

In recent studies involving NAPPA microarrays, extra-well fluorescence is used as a key measure for identifying disease biomarkers because there is evidence to support that it is better correlated with strong antibody responses than statistical analysis involving intraspot intensity. Because this feature is not well quantified by traditional image analysis software, identification and quantification of extra-well fluorescence is performed manually, which is both time-consuming and highly susceptible to variation between raters. A system that could automate this task efficiently and effectively would greatly improve the process of data acquisition in microarray studies, thereby accelerating the discovery of disease biomarkers. In this study, we experimented with different machine learning methods, as well as novel heuristics, for identifying spots exhibiting extra-well fluorescence (rings) in microarray images and assigning each ring a grade of 1-5 based on its intensity and morphology. The sensitivity of our final system for identifying rings was found to be 72% at 99% specificity and 98% at 92% specificity. Our system performs this task significantly faster than a human, while maintaining high performance, and therefore represents a valuable tool for microarray image analysis.

TOXICOGENOMIC ANALYSIS INCORPORATING OPERON-TRANSCRIPTIONAL COUPLING AND TOXICANT CONCENTRATRION-EXPRESSION RESPONSE: Analysis of MX-Treated Salmonella

EPA Science Inventory

What is the study? This study is the first to use microarray analysis in the Ames strains of Salmonella. The microarray chips were custom-designed for this study and are not commercially available, and we evaluated the well-studied drinking water mutagen, MX. Because much inform...

Employing image processing techniques for cancer detection using microarray images.

PubMed

Dehghan Khalilabad, Nastaran; Hassanpour, Hamid

2017-02-01

Microarray technology is a powerful genomic tool for simultaneously studying and analyzing the behavior of thousands of genes. The analysis of images obtained from this technology plays a critical role in the detection and treatment of diseases. The aim of the current study is to develop an automated system for analyzing data from microarray images in order to detect cancerous cases. The proposed system consists of three main phases, namely image processing, data mining, and the detection of the disease. The image processing phase performs operations such as refining image rotation, gridding (locating genes) and extracting raw data from images the data mining includes normalizing the extracted data and selecting the more effective genes. Finally, via the extracted data, cancerous cell is recognized. To evaluate the performance of the proposed system, microarray database is employed which includes Breast cancer, Myeloid Leukemia and Lymphomas from the Stanford Microarray Database. The results indicate that the proposed system is able to identify the type of cancer from the data set with an accuracy of 95.45%, 94.11%, and 100%, respectively. Copyright © 2017 Elsevier Ltd. All rights reserved.

APPLICATION OF CDNA MICROARRAY TO THE STUDY OF ARSENIC TOXICOLOGY AND CARCINOGENESIS

EPA Science Inventory

Arsenic (As) is a common environmental toxicant and known human carcinogen. Epidemiological studies link As exposure to various disorders and cancers. However, the molecular mechanisms for As toxicity and carcinogenicity are not completely known. The cDNA microarray, a high-th...

A Follow-Up of the Multicenter Collaborative Study on HIV-1 Drug Resistance and Tropism Testing Using 454 Ultra Deep Pyrosequencing

PubMed Central

St. John, Elizabeth P.; Simen, Birgitte B.; Turenchalk, Gregory S.; Braverman, Michael S.; Abbate, Isabella; Aerssens, Jeroen; Bouchez, Olivier; Gabriel, Christian; Izopet, Jacques; Meixenberger, Karolin; Di Giallonardo, Francesca; Schlapbach, Ralph; Paredes, Roger; Sakwa, James; Schmitz-Agheguian, Gudrun G.; Thielen, Alexander; Victor, Martin

2016-01-01

Background Ultra deep sequencing is of increasing use not only in research but also in diagnostics. For implementation of ultra deep sequencing assays in clinical laboratories for routine diagnostics, intra- and inter-laboratory testing are of the utmost importance. Methods A multicenter study was conducted to validate an updated assay design for 454 Life Sciences’ GS FLX Titanium system targeting protease/reverse transcriptase (RTP) and env (V3) regions to identify HIV-1 drug-resistance mutations and determine co-receptor use with high sensitivity. The study included 30 HIV-1 subtype B and 6 subtype non-B samples with viral titers (VT) of 3,940–447,400 copies/mL, two dilution series (52,129–1,340 and 25,130–734 copies/mL), and triplicate samples. Amplicons spanning PR codons 10–99, RT codons 1–251 and the entire V3 region were generated using barcoded primers. Analysis was performed using the GS Amplicon Variant Analyzer and geno2pheno for tropism. For comparison, population sequencing was performed using the ViroSeq HIV-1 genotyping system. Results The median sequencing depth across the 11 sites was 1,829 reads per position for RTP (IQR 592–3,488) and 2,410 for V3 (IQR 786–3,695). 10 preselected drug resistant variants were measured across sites and showed high inter-laboratory correlation across all sites with data (P<0.001). The triplicate samples of a plasmid mixture confirmed the high inter-laboratory consistency (mean% ± stdev: 4.6 ±0.5, 4.8 ±0.4, 4.9 ±0.3) and revealed good intra-laboratory consistency (mean% range ± stdev range: 4.2–5.2 ± 0.04–0.65). In the two dilutions series, no variants >20% were missed, variants 2–10% were detected at most sites (even at low VT), and variants 1–2% were detected by some sites. All mutations detected by population sequencing were also detected by UDS. Conclusions This assay design results in an accurate and reproducible approach to analyze HIV-1 mutant spectra, even at variant frequencies well below those routinely detectable by population sequencing. PMID:26756901

Multicenter Evaluation of Epidemiological Typing of Methicillin-Resistant Staphylococcus aureus Strains by Repetitive-Element PCR Analysis

PubMed Central

Deplano, Ariane; Schuermans, Annette; Van Eldere, Johan; Witte, Wolfgang; Meugnier, Hèléne; Etienne, Jerome; Grundmann, Hajo; Jonas, Daniel; Noordhoek, Gerda T.; Dijkstra, Jolanda; van Belkum, Alex; van Leeuwen, Willem; Tassios, Panayotis T.; Legakis, Nicholas J.; van der Zee, Anneke; Bergmans, Anneke; Blanc, Dominique S.; Tenover, Fred C.; Cookson, Barry C.; O'Neil, Gael; Struelens, Marc J.

2000-01-01

Rapid and efficient epidemiologic typing systems would be useful to monitor transmission of methicillin-resistant Staphylococcus aureus (MRSA) at both local and interregional levels. To evaluate the intralaboratory performance and interlaboratory reproducibility of three recently developed repeat-element PCR (rep-PCR) methods for the typing of MRSA, 50 MRSA strains characterized by pulsed-field gel electrophoresis (PFGE) (SmaI) analysis and epidemiological data were blindly typed by inter-IS256, 16S-23S ribosomal DNA (rDNA), and MP3 PCR in 12 laboratories in eight countries using standard reagents and protocols. Performance of typing was defined by reproducibility (R), discriminatory power (D), and agreement with PFGE analysis. Interlaboratory reproducibility of pattern and type classification was assessed visually and using gel analysis software. Each typing method showed a different performance level in each center. In the center performing best with each method, inter-IS256 PCR typing achieved R = 100% and D = 100%; 16S-23S rDNA PCR, R = 100% and D = 82%; and MP3 PCR, R = 80% and D = 83%. Concordance between rep-PCR type and PFGE type ranged by center: 70 to 90% for inter-IS256 PCR, 44 to 57% for 16S-23S rDNA PCR, and 53 to 54% for MP3 PCR analysis. In conclusion, the performance of inter-IS256 PCR typing was similar to that of PFGE analysis in some but not all centers, whereas other rep-PCR protocols showed lower discrimination and intralaboratory reproducibility. None of these assays, however, was sufficiently reproducible for interlaboratory exchange of data. PMID:11015358

Identification of differentially expressed genes and false discovery rate in microarray studies.

PubMed

Gusnanto, Arief; Calza, Stefano; Pawitan, Yudi

2007-04-01

To highlight the development in microarray data analysis for the identification of differentially expressed genes, particularly via control of false discovery rate. The emergence of high-throughput technology such as microarrays raises two fundamental statistical issues: multiplicity and sensitivity. We focus on the biological problem of identifying differentially expressed genes. First, multiplicity arises due to testing tens of thousands of hypotheses, rendering the standard P value meaningless. Second, known optimal single-test procedures such as the t-test perform poorly in the context of highly multiple tests. The standard approach of dealing with multiplicity is too conservative in the microarray context. The false discovery rate concept is fast becoming the key statistical assessment tool replacing the P value. We review the false discovery rate approach and argue that it is more sensible for microarray data. We also discuss some methods to take into account additional information from the microarrays to improve the false discovery rate. There is growing consensus on how to analyse microarray data using the false discovery rate framework in place of the classical P value. Further research is needed on the preprocessing of the raw data, such as the normalization step and filtering, and on finding the most sensitive test procedure.

Systematic spatial bias in DNA microarray hybridization is caused by probe spot position-dependent variability in lateral diffusion.

PubMed

Steger, Doris; Berry, David; Haider, Susanne; Horn, Matthias; Wagner, Michael; Stocker, Roman; Loy, Alexander

2011-01-01

The hybridization of nucleic acid targets with surface-immobilized probes is a widely used assay for the parallel detection of multiple targets in medical and biological research. Despite its widespread application, DNA microarray technology still suffers from several biases and lack of reproducibility, stemming in part from an incomplete understanding of the processes governing surface hybridization. In particular, non-random spatial variations within individual microarray hybridizations are often observed, but the mechanisms underpinning this positional bias remain incompletely explained. This study identifies and rationalizes a systematic spatial bias in the intensity of surface hybridization, characterized by markedly increased signal intensity of spots located at the boundaries of the spotted areas of the microarray slide. Combining observations from a simplified single-probe block array format with predictions from a mathematical model, the mechanism responsible for this bias is found to be a position-dependent variation in lateral diffusion of target molecules. Numerical simulations reveal a strong influence of microarray well geometry on the spatial bias. Reciprocal adjustment of the size of the microarray hybridization chamber to the area of surface-bound probes is a simple and effective measure to minimize or eliminate the diffusion-based bias, resulting in increased uniformity and accuracy of quantitative DNA microarray hybridization.

Systematic Spatial Bias in DNA Microarray Hybridization Is Caused by Probe Spot Position-Dependent Variability in Lateral Diffusion

PubMed Central

Haider, Susanne; Horn, Matthias; Wagner, Michael; Stocker, Roman; Loy, Alexander

2011-01-01

Background The hybridization of nucleic acid targets with surface-immobilized probes is a widely used assay for the parallel detection of multiple targets in medical and biological research. Despite its widespread application, DNA microarray technology still suffers from several biases and lack of reproducibility, stemming in part from an incomplete understanding of the processes governing surface hybridization. In particular, non-random spatial variations within individual microarray hybridizations are often observed, but the mechanisms underpinning this positional bias remain incompletely explained. Methodology/Principal Findings This study identifies and rationalizes a systematic spatial bias in the intensity of surface hybridization, characterized by markedly increased signal intensity of spots located at the boundaries of the spotted areas of the microarray slide. Combining observations from a simplified single-probe block array format with predictions from a mathematical model, the mechanism responsible for this bias is found to be a position-dependent variation in lateral diffusion of target molecules. Numerical simulations reveal a strong influence of microarray well geometry on the spatial bias. Conclusions Reciprocal adjustment of the size of the microarray hybridization chamber to the area of surface-bound probes is a simple and effective measure to minimize or eliminate the diffusion-based bias, resulting in increased uniformity and accuracy of quantitative DNA microarray hybridization. PMID:21858215

Crossword: A Fully Automated Algorithm for the Segmentation and Quality Control of Protein Microarray Images

PubMed Central

2015-01-01

Biological assays formatted as microarrays have become a critical tool for the generation of the comprehensive data sets required for systems-level understanding of biological processes. Manual annotation of data extracted from images of microarrays, however, remains a significant bottleneck, particularly for protein microarrays due to the sensitivity of this technology to weak artifact signal. In order to automate the extraction and curation of data from protein microarrays, we describe an algorithm called Crossword that logically combines information from multiple approaches to fully automate microarray segmentation. Automated artifact removal is also accomplished by segregating structured pixels from the background noise using iterative clustering and pixel connectivity. Correlation of the location of structured pixels across image channels is used to identify and remove artifact pixels from the image prior to data extraction. This component improves the accuracy of data sets while reducing the requirement for time-consuming visual inspection of the data. Crossword enables a fully automated protocol that is robust to significant spatial and intensity aberrations. Overall, the average amount of user intervention is reduced by an order of magnitude and the data quality is increased through artifact removal and reduced user variability. The increase in throughput should aid the further implementation of microarray technologies in clinical studies. PMID:24417579

The detection and differentiation of canine respiratory pathogens using oligonucleotide microarrays.

PubMed

Wang, Lih-Chiann; Kuo, Ya-Ting; Chueh, Ling-Ling; Huang, Dean; Lin, Jiunn-Horng

2017-05-01

Canine respiratory diseases are commonly seen in dogs along with co-infections with multiple respiratory pathogens, including viruses and bacteria. Virus infections in even vaccinated dogs were also reported. The clinical signs caused by different respiratory etiological agents are similar, which makes differential diagnosis imperative. An oligonucleotide microarray system was developed in this study. The wild type and vaccine strains of canine distemper virus (CDV), influenza virus, canine herpesvirus (CHV), Bordetella bronchiseptica and Mycoplasma cynos were detected and differentiated simultaneously on a microarray chip. The detection limit is 10, 10, 100, 50 and 50 copy numbers for CDV, influenza virus, CHV, B. bronchiseptica and M. cynos, respectively. The clinical test results of nasal swab samples showed that the microarray had remarkably better efficacy than the multiplex PCR-agarose gel method. The positive detection rate of microarray and agarose gel was 59.0% (n=33) and 41.1% (n=23) among the 56 samples, respectively. CDV vaccine strain and pathogen co-infections were further demonstrated by the microarray but not by the multiplex PCR-agarose gel. The oligonucleotide microarray provides a highly efficient diagnosis alternative that could be applied to clinical usage, greatly assisting in disease therapy and control. Copyright © 2017 Elsevier B.V. All rights reserved.

Construction of a cDNA microarray derived from the ascidian Ciona intestinalis.

PubMed

Azumi, Kaoru; Takahashi, Hiroki; Miki, Yasufumi; Fujie, Manabu; Usami, Takeshi; Ishikawa, Hisayoshi; Kitayama, Atsusi; Satou, Yutaka; Ueno, Naoto; Satoh, Nori

2003-10-01

A cDNA microarray was constructed from a basal chordate, the ascidian Ciona intestinalis. The draft genome of Ciona has been read and inferred to contain approximately 16,000 protein-coding genes, and cDNAs for transcripts of 13,464 genes have been characterized and compiled as the "Ciona intestinalis Gene Collection Release I". In the present study, we constructed a cDNA microarray of these 13,464 Ciona genes. A preliminary experiment with Cy3- and Cy5-labeled probes showed extensive differential gene expression between fertilized eggs and larvae. In addition, there was a good correlation between results obtained by the present microarray analysis and those from previous EST analyses. This first microarray of a large collection of Ciona intestinalis cDNA clones should facilitate the analysis of global gene expression and gene networks during the embryogenesis of basal chordates.

Improvement in the amine glass platform by bubbling method for a DNA microarray

PubMed Central

Jee, Seung Hyun; Kim, Jong Won; Lee, Ji Hyeong; Yoon, Young Soo

2015-01-01

A glass platform with high sensitivity for sexually transmitted diseases microarray is described here. An amino-silane-based self-assembled monolayer was coated on the surface of a glass platform using a novel bubbling method. The optimized surface of the glass platform had highly uniform surface modifications using this method, as well as improved hybridization properties with capture probes in the DNA microarray. On the basis of these results, the improved glass platform serves as a highly reliable and optimal material for the DNA microarray. Moreover, in this study, we demonstrated that our glass platform, manufactured by utilizing the bubbling method, had higher uniformity, shorter processing time, lower background signal, and higher spot signal than the platforms manufactured by the general dipping method. The DNA microarray manufactured with a glass platform prepared using bubbling method can be used as a clinical diagnostic tool. PMID:26468293

Improvement in the amine glass platform by bubbling method for a DNA microarray.

PubMed

Jee, Seung Hyun; Kim, Jong Won; Lee, Ji Hyeong; Yoon, Young Soo

2015-01-01

A glass platform with high sensitivity for sexually transmitted diseases microarray is described here. An amino-silane-based self-assembled monolayer was coated on the surface of a glass platform using a novel bubbling method. The optimized surface of the glass platform had highly uniform surface modifications using this method, as well as improved hybridization properties with capture probes in the DNA microarray. On the basis of these results, the improved glass platform serves as a highly reliable and optimal material for the DNA microarray. Moreover, in this study, we demonstrated that our glass platform, manufactured by utilizing the bubbling method, had higher uniformity, shorter processing time, lower background signal, and higher spot signal than the platforms manufactured by the general dipping method. The DNA microarray manufactured with a glass platform prepared using bubbling method can be used as a clinical diagnostic tool.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gentry, T.; Schadt, C.; Zhou, J.

Microarray technology has the unparalleled potential tosimultaneously determine the dynamics and/or activities of most, if notall, of the microbial populations in complex environments such as soilsand sediments. Researchers have developed several types of arrays thatcharacterize the microbial populations in these samples based on theirphylogenetic relatedness or functional genomic content. Several recentstudies have used these microarrays to investigate ecological issues;however, most have only analyzed a limited number of samples withrelatively few experiments utilizing the full high-throughput potentialof microarray analysis. This is due in part to the unique analyticalchallenges that these samples present with regard to sensitivity,specificity, quantitation, and data analysis. Thismore » review discussesspecific applications of microarrays to microbial ecology research alongwith some of the latest studies addressing the difficulties encounteredduring analysis of complex microbial communities within environmentalsamples. With continued development, microarray technology may ultimatelyachieve its potential for comprehensive, high-throughput characterizationof microbial populations in near real-time.« less

Development of a rapid microarray-based DNA subtyping assay for the alleles of Shiga toxins 1 and 2 of Escherichia coli.

PubMed

Geue, Lutz; Stieber, Bettina; Monecke, Stefan; Engelmann, Ines; Gunzer, Florian; Slickers, Peter; Braun, Sascha D; Ehricht, Ralf

2014-08-01

In this study, we developed a new rapid, economic, and automated microarray-based genotyping test for the standardized subtyping of Shiga toxins 1 and 2 of Escherichia coli. The microarrays from Alere Technologies can be used in two different formats, the ArrayTube and the ArrayStrip (which enables high-throughput testing in a 96-well format). One microarray chip harbors all the gene sequences necessary to distinguish between all Stx subtypes, facilitating the identification of single and multiple subtypes within a single isolate in one experiment. Specific software was developed to automatically analyze all data obtained from the microarray. The assay was validated with 21 Shiga toxin-producing E. coli (STEC) reference strains that were previously tested by the complete set of conventional subtyping PCRs. The microarray results showed 100% concordance with the PCR results. Essentially identical results were detected when the standard DNA extraction method was replaced by a time-saving heat lysis protocol. For further validation of the microarray, we identified the Stx subtypes or combinations of the subtypes in 446 STEC field isolates of human and animal origin. In summary, this oligonucleotide array represents an excellent diagnostic tool that provides some advantages over standard PCR-based subtyping. The number of the spotted probes on the microarrays can be increased by additional probes, such as for novel alleles, species markers, or resistance genes, should the need arise. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Standardization of reflectance measurements in dispersed organic matter: results of an exercise to improve interlaboratory agreement

USGS Publications Warehouse

Hackley, Paul C.; Araujo, Carla Viviane; Borrego, Angeles G.; Bouzinos, Antonis; Cardott, Brian; Cook, Alan C.; Eble, Cortland; Flores, Deolinda; Gentzis, Thomas; Gonçalves, Paula Alexandra; Filho, João Graciano Mendonça; Hámor-Vidó, Mária; Jelonek, Iwona; Kommeren, Kees; Knowles, Wayne; Kus, Jolanta; Mastalerz, Maria; Menezes, Taíssa Rêgo; Newman, Jane; Pawlewicz, Mark; Pickel, Walter; Potter, Judith; Ranasinghe, Paddy; Read, Harold; Reyes, Julito; Rodriguez, Genaro De La Rosa; de Souza, Igor Viegas Alves Fernandes; Suarez-Ruiz, Isabel; Sýkorová, Ivana; Valentine, Brett J.

2015-01-01

Vitrinite reflectance generally is considered the most robust thermal maturity parameter available for application to hydrocarbon exploration and petroleum system evaluation. However, until 2011 there was no standardized methodology available to provide guidelines for vitrinite reflectance measurements in shale. Efforts to correct this deficiency resulted in publication of ASTM D7708: Standard test method for microscopical determination of the reflectance of vitrinite dispersed in sedimentary rocks. In 2012-2013, an interlaboratory exercise was conducted to establish precision limits for the D7708 measurement technique. Six samples, representing a wide variety of shale, were tested in duplicate by 28 analysts in 22 laboratories from 14 countries. Samples ranged from immature to overmature (0.31-1.53% Ro), from organic-lean to organic-rich (1-22 wt.% total organic carbon), and contained Type I (lacustrine), Type II (marine), and Type III (terrestrial) kerogens. Repeatability limits (maximum difference between valid repetitive results from same operator, same conditions) ranged from 0.03-0.11% absolute reflectance, whereas reproducibility limits (maximum difference between valid results obtained on same test material by different operators, different laboratories) ranged from 0.12-0.54% absolute reflectance. Repeatability and reproducibility limits degraded consistently with increasing maturity and decreasing organic content. However, samples with terrestrial kerogens (Type III) fell off this trend, showing improved levels of reproducibility due to higher vitrinite content and improved ease of identification. Operators did not consistently meet the reporting requirements of the test method, indicating that a common reporting template is required to improve data quality. The most difficult problem encountered was the petrographic distinction of solid bitumens and low-reflecting inert macerals from vitrinite when vitrinite occurred with reflectance ranges overlapping the other components. Discussion among participants suggested this problem could not be easily corrected via kerogen concentration or solvent extraction and is related to operator training and background. No statistical difference in mean reflectance was identified between participants reporting bitumen reflectance vs. vitrinite reflectance vs. a mixture of bitumen and vitrinite reflectance values, suggesting empirical conversion schemes should be treated with caution. Analysis of reproducibility limits obtained during this exercise in comparison to reproducibility limits from historical interlaboratory exercises suggests use of a common methodology (D7708) improves interlaboratory precision. Future work will investigate opportunities to improve reproducibility in high maturity, organic-lean shale varieties.

A Review of Positive Ion Sensitivities for the SIMS Analysis of CMT

DTIC Science & Technology

1991-05-01

microprobe. Inter-laboratory exercises organised by NRL using standardised glasses and steels’ s showed considerable agreement usually within a factor...would be sufficient oxygen to convert all the remaining matrix atoms to oxides, TeO2 and CdO. Any general theory of the lonisation of sputtered particles...Eggert equation which works well for many other matrices, such as metals, glasses and ceramics. Despite decades of basic studies there is still no

Accuracy of trace element determinations in alternate fuels

NASA Technical Reports Server (NTRS)

Greenbauer-Seng, L. A.

1980-01-01

NASA-Lewis Research Center's work on accurate measurement of trace level of metals in various fuels is presented. The differences between laboratories and between analytical techniques especially for concentrations below 10 ppm, are discussed, detailing the Atomic Absorption Spectrometry (AAS) and DC Arc Emission Spectrometry (dc arc) techniques used by NASA-Lewis. Also presented is the design of an Interlaboratory Study which is considering the following factors: laboratory, analytical technique, fuel type, concentration and ashing additive.

The efficacy of microarray screening for autosomal recessive retinitis pigmentosa in routine clinical practice

PubMed Central

van Huet, Ramon A. C.; Pierrache, Laurence H.M.; Meester-Smoor, Magda A.; Klaver, Caroline C.W.; van den Born, L. Ingeborgh; Hoyng, Carel B.; de Wijs, Ilse J.; Collin, Rob W. J.; Hoefsloot, Lies H.

2015-01-01

Purpose To determine the efficacy of multiple versions of a commercially available arrayed primer extension (APEX) microarray chip for autosomal recessive retinitis pigmentosa (arRP). Methods We included 250 probands suspected of arRP who were genetically analyzed with the APEX microarray between January 2008 and November 2013. The mode of inheritance had to be autosomal recessive according to the pedigree (including isolated cases). If the microarray identified a heterozygous mutation, we performed Sanger sequencing of exons and exon–intron boundaries of that specific gene. The efficacy of this microarray chip with the additional Sanger sequencing approach was determined by the percentage of patients that received a molecular diagnosis. We also collected data from genetic tests other than the APEX analysis for arRP to provide a detailed description of the molecular diagnoses in our study cohort. Results The APEX microarray chip for arRP identified the molecular diagnosis in 21 (8.5%) of the patients in our cohort. Additional Sanger sequencing yielded a second mutation in 17 patients (6.8%), thereby establishing the molecular diagnosis. In total, 38 patients (15.2%) received a molecular diagnosis after analysis using the microarray and additional Sanger sequencing approach. Further genetic analyses after a negative result of the arRP microarray (n = 107) resulted in a molecular diagnosis of arRP (n = 23), autosomal dominant RP (n = 5), X-linked RP (n = 2), and choroideremia (n = 1). Conclusions The efficacy of the commercially available APEX microarray chips for arRP appears to be low, most likely caused by the limitations of this technique and the genetic and allelic heterogeneity of RP. Diagnostic yields up to 40% have been reported for next-generation sequencing (NGS) techniques that, as expected, thereby outperform targeted APEX analysis. PMID:25999674

OECD validation study to assess intra- and inter-laboratory reproducibility of the zebrafish embryo toxicity test for acute aquatic toxicity testing.

PubMed

Busquet, François; Strecker, Ruben; Rawlings, Jane M; Belanger, Scott E; Braunbeck, Thomas; Carr, Gregory J; Cenijn, Peter; Fochtman, Przemyslaw; Gourmelon, Anne; Hübler, Nicole; Kleensang, André; Knöbel, Melanie; Kussatz, Carola; Legler, Juliette; Lillicrap, Adam; Martínez-Jerónimo, Fernando; Polleichtner, Christian; Rzodeczko, Helena; Salinas, Edward; Schneider, Katharina E; Scholz, Stefan; van den Brandhof, Evert-Jan; van der Ven, Leo T M; Walter-Rohde, Susanne; Weigt, Stefan; Witters, Hilda; Halder, Marlies

2014-08-01

The OECD validation study of the zebrafish embryo acute toxicity test (ZFET) for acute aquatic toxicity testing evaluated the ZFET reproducibility by testing 20 chemicals at 5 different concentrations in 3 independent runs in at least 3 laboratories. Stock solutions and test concentrations were analytically confirmed for 11 chemicals. Newly fertilised zebrafish eggs (20/concentration and control) were exposed for 96h to chemicals. Four apical endpoints were recorded daily as indicators of acute lethality: coagulation of the embryo, lack of somite formation, non-detachment of the tail bud from the yolk sac and lack of heartbeat. Results (LC50 values for 48/96h exposure) show that the ZFET is a robust method with a good intra- and inter-laboratory reproducibility (CV<30%) for most chemicals and laboratories. The reproducibility was lower (CV>30%) for some very toxic or volatile chemicals, and chemicals tested close to their limit of solubility. The ZFET is now available as OECD Test Guideline 236. Considering the high predictive capacity of the ZFET demonstrated by Belanger et al. (2013) in their retrospective analysis of acute fish toxicity and fish embryo acute toxicity data, the ZFET is ready to be considered for acute fish toxicity for regulatory purposes. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Interlaboratory studies with the Chinese hamster V79 cell metabolic cooperation assay to detect tumor-promoting agents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bohrman, J.S.; Burg, J.R.; Elmore, E.

1988-01-01

Three laboratories participated in an interlaboratory study to evaluate the usefulness of the Chinese hamster V79 cell metabolic cooperation assay to predict the tumor-promoting activity of selected chemical. Twenty-three chemicals of different chemical structures (phorbol esters, barbiturates, phenols, artificial sweeteners, alkanes, and peroxides) were chosen for testing based on in vivo promotion activities, as reported in the literature. Assay protocols and materials were standardized, and the chemicals were coded to facilitate unbiased evaluation. A chemical was tested only once in each laboratory, with one of the three laboratories testing only 15 out of 23 chemicals. Dunnett's test was used formore » statistical analysis. Chemicals were scored as positive (at least two concentration levels statistically different than control), equivocal (only one concentration statistically different), or negative. For 15 chemicals tested in all three laboratories, there was complete agreement among the laboratories for nine chemicals. For the 23 chemicals tested in only two laboratories, there was agreement on 16 chemicals. With the exception of the peroxides and alkanes, the metabolic cooperation data were in general agreement with in vivo data. However, an overall evaluation of the V79 cell system for predicting in vivo promotion activity was difficult because of the organ specificity of certain chemicals and/or the limited number of adequately tested nonpromoting chemicals.« less

Interlaboratory study of free cyanide methods compared to total cyanide measurements and the effect of preservation with sodium hydroxide for secondary- and tertiary-treated waste water samples.

PubMed

Stanley, Brett J; Antonio, Karen

2012-11-01

Several methods exist for the measurement of cyanide levels in treated wastewater,typically requiring preservation of the sample with sodium hydroxide to minimize loss of hydrogen cyanide gas (HCN). Recent reports have shown that cyanide levels may increase with chlorination or preservation. In this study, three flow injection analysis methods involving colorimetric and amperometric detection were compared within one laboratory, as well as across separate laboratories and equipment. Split wastewater samples from eight facilities and three different sampling periods were tested. An interlaboratory confidence interval of 3.5 ppb was calculated compared with the intralaboratory reporting limit of 2 ppb. The results show that free cyanide measurements are not statistically different than total cyanide levels. An artificial increase in cyanide level is observed with all methods for preserved samples relative to nonpreserved samples, with an average increase of 2.3 ppb. The possible loss of cyanide without preservation is shown to be statistically insignificant if properly stored up to 48 hours. The cyanide increase with preservation is further substantiated with the method of standard additions and is not a matrix interference. The increase appears to be correlated with the amount of cyanide observed without preservation, which appears to be greater in those facilities that disinfect their wastewater with chlorine, followed by dechlorination with sodium bisulfite.

Evaluation of Cardiac Toxicity Biomarkers in Rats from Different Laboratories

PubMed Central

Kim, Kyuri; Chini, Naseem; Fairchild, David G.; Engle, Steven K.; Reagan, William J.; Summers, Sandra D.; Mirsalis, Jon C.

2016-01-01

There is a great need for improved diagnostic and prognostic accuracy of potential cardiac toxicity in drug development. This study reports the evaluation of several commercially available biomarker kits by three institutions (SRI, Eli Lilly and Pfizer) for the discrimination between myocardial degeneration/necrosis and cardiac hypertrophy as well as the assessment of the inter-laboratory and inter-platform variation in results. Serum concentrations of natriuretic peptides (NT-proANP, NT-proBNP), cardiac and skeletal troponins (cTnI, cTnT, sTnI), myosin light chain 3 (Myl3) and fatty acid binding protein 3 (FABP3) were assessed in rats treated with minoxidil and isoproterenol. Minoxidil caused increased heart-to-body weight ratios and prominent elevations in NT-proANP and NT-proBNP concentrations detected at 24 hr postdose without elevation in troponins, Myl3 or FABP3 and with no abnormal histopathological findings. Isoproterenol caused ventricular leukocyte infiltration, myocyte fibrosis and necrosis with increased concentrations of the natriuretic peptides, cardiac troponins and Myl3. These results reinforce the advantages of a multi-marker strategy in elucidating the underlying cause of cardiac insult and detecting myocardial tissue damage at 24 hr post-treatment. The inter-laboratory and inter-platform comparison analyses also showed that the data obtained from different laboratories and platforms are highly correlated and reproducible, making these biomarkers widely applicable in preclinical studies. PMID:27638646

On the stability of a variety of organic photovoltaic devices by IPCE and in situ IPCE analyses--the ISOS-3 inter-laboratory collaboration.

PubMed

Teran-Escobar, Gerardo; Tanenbaum, David M; Voroshazi, Eszter; Hermenau, Martin; Norrman, Kion; Lloyd, Matthew T; Galagan, Yulia; Zimmermann, Birger; Hösel, Markus; Dam, Henrik F; Jørgensen, Mikkel; Gevorgyan, Suren; Kudret, Suleyman; Maes, Wouter; Lutsen, Laurence; Vanderzande, Dirk; Würfel, Uli; Andriessen, Ronn; Rösch, Roland; Hoppe, Harald; Rivaton, Agnès; Uzunoğlu, Gülşah Y; Germack, David; Andreasen, Birgitta; Madsen, Morten V; Bundgaard, Eva; Krebs, Frederik C; Lira-Cantu, Monica

2012-09-07

This work is part of the inter-laboratory collaboration to study the stability of seven distinct sets of state-of-the-art organic photovoltaic (OPV) devices prepared by leading research laboratories. All devices have been shipped to and degraded at RISØ-DTU up to 1830 hours in accordance with established ISOS-3 protocols under defined illumination conditions. In this work, we apply the Incident Photon-to-Electron Conversion Efficiency (IPCE) and the in situ IPCE techniques to determine the relation between solar cell performance and solar cell stability. Different ageing conditions were considered: accelerated full sun simulation, low level indoor fluorescent lighting and dark storage. The devices were also monitored under conditions of ambient and inert (N(2)) atmospheres, which allows for the identification of the solar cell materials more susceptible to degradation by ambient air (oxygen and moisture). The different OPVs configurations permitted the study of the intrinsic stability of the devices depending on: two different ITO-replacement alternatives, two different hole extraction layers (PEDOT:PSS and MoO(3)), and two different P3HT-based polymers. The response of un-encapsulated devices to ambient atmosphere offered insight into the importance of moisture in solar cell performance. Our results demonstrate that the IPCE and the in situ IPCE techniques are valuable analytical methods to understand device degradation and solar cell lifetime.

Size characterization of airborne SiO2 nanoparticles with on-line and off-line measurement techniques: an interlaboratory comparison study

NASA Astrophysics Data System (ADS)

Motzkus, C.; Macé, T.; Gaie-Levrel, F.; Ducourtieux, S.; Delvallee, A.; Dirscherl, K.; Hodoroaba, V.-D.; Popov, I.; Popov, O.; Kuselman, I.; Takahata, K.; Ehara, K.; Ausset, P.; Maillé, M.; Michielsen, N.; Bondiguel, S.; Gensdarmes, F.; Morawska, L.; Johnson, G. R.; Faghihi, E. M.; Kim, C. S.; Kim, Y. H.; Chu, M. C.; Guardado, J. A.; Salas, A.; Capannelli, G.; Costa, C.; Bostrom, T.; Jämting, Å. K.; Lawn, M. A.; Adlem, L.; Vaslin-Reimann, S.

2013-10-01

Results of an interlaboratory comparison on size characterization of SiO2 airborne nanoparticles using on-line and off-line measurement techniques are discussed. This study was performed in the framework of Technical Working Area (TWA) 34—"Properties of Nanoparticle Populations" of the Versailles Project on Advanced Materials and Standards (VAMAS) in the project no. 3 "Techniques for characterizing size distribution of airborne nanoparticles". Two types of nano-aerosols, consisting of (1) one population of nanoparticles with a mean diameter between 30.3 and 39.0 nm and (2) two populations of non-agglomerated nanoparticles with mean diameters between, respectively, 36.2-46.6 nm and 80.2-89.8 nm, were generated for characterization measurements. Scanning mobility particle size spectrometers (SMPS) were used for on-line measurements of size distributions of the produced nano-aerosols. Transmission electron microscopy, scanning electron microscopy, and atomic force microscopy were used as off-line measurement techniques for nanoparticles characterization. Samples were deposited on appropriate supports such as grids, filters, and mica plates by electrostatic precipitation and a filtration technique using SMPS controlled generation upstream. The results of the main size distribution parameters (mean and mode diameters), obtained from several laboratories, were compared based on metrological approaches including metrological traceability, calibration, and evaluation of the measurement uncertainty. Internationally harmonized measurement procedures for airborne SiO2 nanoparticles characterization are proposed.

A highly sensitive and specific method for the screening detection of genetically modified organisms based on digital PCR without pretreatment

PubMed Central

Fu, Wei; Zhu, Pengyu; Wang, Chenguang; Huang, Kunlun; Du, Zhixin; Tian, Wenying; Wang, Qin; Wang, Huiyu; Xu, Wentao; Zhu, Shuifang

2015-01-01

Digital PCR has developed rapidly since it was first reported in the 1990s. It was recently reported that an improved method facilitated the detection of genetically modified organisms (GMOs). However, to use this improved method, the samples must be pretreated, which could introduce inaccuracy into the results. In our study, we explored a pretreatment-free digital PCR detection method for the screening for GMOs. We chose the CaMV35s promoter and the NOS terminator as the templates in our assay. To determine the specificity of our method, 9 events of GMOs were collected, including MON810, MON863, TC1507, MIR604, MIR162, GA21, T25, NK603 and Bt176. Moreover, the sensitivity, intra-laboratory and inter-laboratory reproducibility of our detection method were assessed. The results showed that the limit of detection of our method was 0.1%, which was lower than the labeling threshold level of the EU. The specificity and stability among the 9 events were consistent, respectively. The intra-laboratory and inter-laboratory reproducibility were both good. Finally, the perfect fitness for the detection of eight double-blind samples indicated the good practicability of our method. In conclusion, the method in our study would allow more sensitive, specific and stable screening detection of the GMO content of international trading products. PMID:26239916

A highly sensitive and specific method for the screening detection of genetically modified organisms based on digital PCR without pretreatment.

PubMed

Fu, Wei; Zhu, Pengyu; Wang, Chenguang; Huang, Kunlun; Du, Zhixin; Tian, Wenying; Wang, Qin; Wang, Huiyu; Xu, Wentao; Zhu, Shuifang

2015-08-04

Digital PCR has developed rapidly since it was first reported in the 1990 s. It was recently reported that an improved method facilitated the detection of genetically modified organisms (GMOs). However, to use this improved method, the samples must be pretreated, which could introduce inaccuracy into the results. In our study, we explored a pretreatment-free digital PCR detection method for the screening for GMOs. We chose the CaMV35s promoter and the NOS terminator as the templates in our assay. To determine the specificity of our method, 9 events of GMOs were collected, including MON810, MON863, TC1507, MIR604, MIR162, GA21, T25, NK603 and Bt176. Moreover, the sensitivity, intra-laboratory and inter-laboratory reproducibility of our detection method were assessed. The results showed that the limit of detection of our method was 0.1%, which was lower than the labeling threshold level of the EU. The specificity and stability among the 9 events were consistent, respectively. The intra-laboratory and inter-laboratory reproducibility were both good. Finally, the perfect fitness for the detection of eight double-blind samples indicated the good practicability of our method. In conclusion, the method in our study would allow more sensitive, specific and stable screening detection of the GMO content of international trading products.

Gene expression profiling of whole blood: Comparison of target preparation methods for accurate and reproducible microarray analysis

PubMed Central

Vartanian, Kristina; Slottke, Rachel; Johnstone, Timothy; Casale, Amanda; Planck, Stephen R; Choi, Dongseok; Smith, Justine R; Rosenbaum, James T; Harrington, Christina A

2009-01-01

Background Peripheral blood is an accessible and informative source of transcriptomal information for many human disease and pharmacogenomic studies. While there can be significant advantages to analyzing RNA isolated from whole blood, particularly in clinical studies, the preparation of samples for microarray analysis is complicated by the need to minimize artifacts associated with highly abundant globin RNA transcripts. The impact of globin RNA transcripts on expression profiling data can potentially be reduced by using RNA preparation and labeling methods that remove or block globin RNA during the microarray assay. We compared four different methods for preparing microarray hybridization targets from human whole blood collected in PAXGene tubes. Three of the methods utilized the Affymetrix one-cycle cDNA synthesis/in vitro transcription protocol but varied treatment of input RNA as follows: i. no treatment; ii. treatment with GLOBINclear; or iii. treatment with globin PNA oligos. In the fourth method cDNA targets were prepared with the Ovation amplification and labeling system. Results We find that microarray targets generated with labeling methods that reduce globin mRNA levels or minimize the impact of globin transcripts during hybridization detect more transcripts in the microarray assay compared with the standard Affymetrix method. Comparison of microarray results with quantitative PCR analysis of a panel of genes from the NF-kappa B pathway shows good correlation of transcript measurements produced with all four target preparation methods, although method-specific differences in overall correlation were observed. The impact of freezing blood collected in PAXGene tubes on data reproducibility was also examined. Expression profiles show little or no difference when RNA is extracted from either fresh or frozen blood samples. Conclusion RNA preparation and labeling methods designed to reduce the impact of globin mRNA transcripts can significantly improve the sensitivity of the DNA microarray expression profiling assay for whole blood samples. While blockage of globin transcripts during first strand cDNA synthesis with globin PNAs resulted in the best overall performance in this study, we conclude that selection of a protocol for expression profiling studies in blood should depend on several factors, including implementation requirements of the method and study design. RNA isolated from either freshly collected or frozen blood samples stored in PAXGene tubes can be used without altering gene expression profiles. PMID:19123946

Importing MAGE-ML format microarray data into BioConductor.

PubMed

Durinck, Steffen; Allemeersch, Joke; Carey, Vincent J; Moreau, Yves; De Moor, Bart

2004-12-12

The microarray gene expression markup language (MAGE-ML) is a widely used XML (eXtensible Markup Language) standard for describing and exchanging information about microarray experiments. It can describe microarray designs, microarray experiment designs, gene expression data and data analysis results. We describe RMAGEML, a new Bioconductor package that provides a link between cDNA microarray data stored in MAGE-ML format and the Bioconductor framework for preprocessing, visualization and analysis of microarray experiments. http://www.bioconductor.org. Open Source.

The future of microarray technology: networking the genome search.

PubMed

D'Ambrosio, C; Gatta, L; Bonini, S

2005-10-01

In recent years microarray technology has been increasingly used in both basic and clinical research, providing substantial information for a better understanding of genome-environment interactions responsible for diseases, as well as for their diagnosis and treatment. However, in genomic research using microarray technology there are several unresolved issues, including scientific, ethical and legal issues. Networks of excellence like GA(2)LEN may represent the best approach for teaching, cost reduction, data repositories, and functional studies implementation.

Fiber-optic microarray for simultaneous detection of multiple harmful algal bloom species.

PubMed

Ahn, Soohyoun; Kulis, David M; Erdner, Deana L; Anderson, Donald M; Walt, David R

2006-09-01

Harmful algal blooms (HABs) are a serious threat to coastal resources, causing a variety of impacts on public health, regional economies, and ecosystems. Plankton analysis is a valuable component of many HAB monitoring and research programs, but the diversity of plankton poses a problem in discriminating toxic from nontoxic species using conventional detection methods. Here we describe a sensitive and specific sandwich hybridization assay that combines fiber-optic microarrays with oligonucleotide probes to detect and enumerate the HAB species Alexandrium fundyense, Alexandrium ostenfeldii, and Pseudo-nitzschia australis. Microarrays were prepared by loading oligonucleotide probe-coupled microspheres (diameter, 3 mum) onto the distal ends of chemically etched imaging fiber bundles. Hybridization of target rRNA from HAB cells to immobilized probes on the microspheres was visualized using Cy3-labeled secondary probes in a sandwich-type assay format. We applied these microarrays to the detection and enumeration of HAB cells in both cultured and field samples. Our study demonstrated a detection limit of approximately 5 cells for all three target organisms within 45 min, without a separate amplification step, in both sample types. We also developed a multiplexed microarray to detect the three HAB species simultaneously, which successfully detected the target organisms, alone and in combination, without cross-reactivity. Our study suggests that fiber-optic microarrays can be used for rapid and sensitive detection and potential enumeration of HAB species in the environment.

Development and assessment of whole-genome oligonucleotide microarrays to analyze an anaerobic microbial community and its responses to oxidative stress.

PubMed

Scholten, Johannes C M; Culley, David E; Nie, Lei; Munn, Kyle J; Chow, Lely; Brockman, Fred J; Zhang, Weiwen

2007-06-29

The application of DNA microarray technology to investigate multiple-species microbial communities presents great challenges. In this study, we reported the design and quality assessment of four whole genome oligonucleotide microarrays for two syntroph bacteria, Desulfovibrio vulgaris and Syntrophobacter fumaroxidans, and two archaeal methanogens, Methanosarcina barkeri, and Methanospirillum hungatei, and their application to analyze global gene expression in a four-species microbial community in response to oxidative stress. In order to minimize the possibility of cross-hybridization, cross-genome comparison was performed to assure all probes unique to each genome so that the microarrays could provide species-level resolution. Microarray quality was validated by the good reproducibility of experimental measurements of multiple biological and analytical replicates. This study showed that S. fumaroxidans and M. hungatei responded to the oxidative stress with up-regulation of several genes known to be involved in reactive oxygen species (ROS) detoxification, such as catalase and rubrerythrin in S. fumaroxidans and thioredoxin and heat shock protein Hsp20 in M. hungatei. However, D. vulgaris seemed to be less sensitive to the oxidative stress as a member of a four-species community, since no gene involved in ROS detoxification was up-regulated. Our work demonstrated the successful application of microarrays to a multiple-species microbial community, and our preliminary results indicated that this approach could provide novel insights on the metabolism within microbial communities.

Systematic evaluation of RNA quality, microarray data reliability and pathway analysis in fresh, fresh frozen and formalin-fixed paraffin-embedded tissue samples.

PubMed

Wimmer, Isabella; Tröscher, Anna R; Brunner, Florian; Rubino, Stephen J; Bien, Christian G; Weiner, Howard L; Lassmann, Hans; Bauer, Jan

2018-04-20

Formalin-fixed paraffin-embedded (FFPE) tissues are valuable resources commonly used in pathology. However, formalin fixation modifies nucleic acids challenging the isolation of high-quality RNA for genetic profiling. Here, we assessed feasibility and reliability of microarray studies analysing transcriptome data from fresh, fresh-frozen (FF) and FFPE tissues. We show that reproducible microarray data can be generated from only 2 ng FFPE-derived RNA. For RNA quality assessment, fragment size distribution (DV200) and qPCR proved most suitable. During RNA isolation, extending tissue lysis time to 10 hours reduced high-molecular-weight species, while additional incubation at 70 °C markedly increased RNA yields. Since FF- and FFPE-derived microarrays constitute different data entities, we used indirect measures to investigate gene signal variation and relative gene expression. Whole-genome analyses revealed high concordance rates, while reviewing on single-genes basis showed higher data variation in FFPE than FF arrays. Using an experimental model, gene set enrichment analysis (GSEA) of FFPE-derived microarrays and fresh tissue-derived RNA-Seq datasets yielded similarly affected pathways confirming the applicability of FFPE tissue in global gene expression analysis. Our study provides a workflow comprising RNA isolation, quality assessment and microarray profiling using minimal RNA input, thus enabling hypothesis-generating pathway analyses from limited amounts of precious, pathologically significant FFPE tissues.

Evaluation of gene expression classification studies: factors associated with classification performance.

PubMed

Novianti, Putri W; Roes, Kit C B; Eijkemans, Marinus J C

2014-01-01

Classification methods used in microarray studies for gene expression are diverse in the way they deal with the underlying complexity of the data, as well as in the technique used to build the classification model. The MAQC II study on cancer classification problems has found that performance was affected by factors such as the classification algorithm, cross validation method, number of genes, and gene selection method. In this paper, we study the hypothesis that the disease under study significantly determines which method is optimal, and that additionally sample size, class imbalance, type of medical question (diagnostic, prognostic or treatment response), and microarray platform are potentially influential. A systematic literature review was used to extract the information from 48 published articles on non-cancer microarray classification studies. The impact of the various factors on the reported classification accuracy was analyzed through random-intercept logistic regression. The type of medical question and method of cross validation dominated the explained variation in accuracy among studies, followed by disease category and microarray platform. In total, 42% of the between study variation was explained by all the study specific and problem specific factors that we studied together.

Particle atlas of World Trade Center dust

USGS Publications Warehouse

Lowers, Heather; Meeker, Gregory P.

2005-01-01

The United States Environmental Protection Agency (EPA) has begun a reassessment of the presence of World Trade Center (WTC) dust in residences, public buildings, and office spaces in New York City, New York. Background dust samples collected from residences, public buildings, and office spaces will be analyzed by multiple laboratories for the presence of WTC dust. Other laboratories are currently studying WTC dust for other purposes, such as health effects studies. To assist in inter-laboratory consistency for identification of WTC dust components, this particle atlas of phases in WTC dust has been compiled.

permGPU: Using graphics processing units in RNA microarray association studies.

PubMed

Shterev, Ivo D; Jung, Sin-Ho; George, Stephen L; Owzar, Kouros

2010-06-16

Many analyses of microarray association studies involve permutation, bootstrap resampling and cross-validation, that are ideally formulated as embarrassingly parallel computing problems. Given that these analyses are computationally intensive, scalable approaches that can take advantage of multi-core processor systems need to be developed. We have developed a CUDA based implementation, permGPU, that employs graphics processing units in microarray association studies. We illustrate the performance and applicability of permGPU within the context of permutation resampling for a number of test statistics. An extensive simulation study demonstrates a dramatic increase in performance when using permGPU on an NVIDIA GTX 280 card compared to an optimized C/C++ solution running on a conventional Linux server. permGPU is available as an open-source stand-alone application and as an extension package for the R statistical environment. It provides a dramatic increase in performance for permutation resampling analysis in the context of microarray association studies. The current version offers six test statistics for carrying out permutation resampling analyses for binary, quantitative and censored time-to-event traits.

Microarray Я US: a user-friendly graphical interface to Bioconductor tools that enables accurate microarray data analysis and expedites comprehensive functional analysis of microarray results.

PubMed

Dai, Yilin; Guo, Ling; Li, Meng; Chen, Yi-Bu

2012-06-08

Microarray data analysis presents a significant challenge to researchers who are unable to use the powerful Bioconductor and its numerous tools due to their lack of knowledge of R language. Among the few existing software programs that offer a graphic user interface to Bioconductor packages, none have implemented a comprehensive strategy to address the accuracy and reliability issue of microarray data analysis due to the well known probe design problems associated with many widely used microarray chips. There is also a lack of tools that would expedite the functional analysis of microarray results. We present Microarray Я US, an R-based graphical user interface that implements over a dozen popular Bioconductor packages to offer researchers a streamlined workflow for routine differential microarray expression data analysis without the need to learn R language. In order to enable a more accurate analysis and interpretation of microarray data, we incorporated the latest custom probe re-definition and re-annotation for Affymetrix and Illumina chips. A versatile microarray results output utility tool was also implemented for easy and fast generation of input files for over 20 of the most widely used functional analysis software programs. Coupled with a well-designed user interface, Microarray Я US leverages cutting edge Bioconductor packages for researchers with no knowledge in R language. It also enables a more reliable and accurate microarray data analysis and expedites downstream functional analysis of microarray results.

Versailles Project on Advanced Materials and Standards Interlaboratory Study on Measuring the Thickness and Chemistry of Nanoparticle Coatings Using XPS and LEIS.

PubMed

Belsey, Natalie A; Cant, David J H; Minelli, Caterina; Araujo, Joyce R; Bock, Bernd; Brüner, Philipp; Castner, David G; Ceccone, Giacomo; Counsell, Jonathan D P; Dietrich, Paul M; Engelhard, Mark H; Fearn, Sarah; Galhardo, Carlos E; Kalbe, Henryk; Won Kim, Jeong; Lartundo-Rojas, Luis; Luftman, Henry S; Nunney, Tim S; Pseiner, Johannes; Smith, Emily F; Spampinato, Valentina; Sturm, Jacobus M; Thomas, Andrew G; Treacy, Jon P W; Veith, Lothar; Wagstaffe, Michael; Wang, Hai; Wang, Meiling; Wang, Yung-Chen; Werner, Wolfgang; Yang, Li; Shard, Alexander G

2016-10-27

We report the results of a VAMAS (Versailles Project on Advanced Materials and Standards) inter-laboratory study on the measurement of the shell thickness and chemistry of nanoparticle coatings. Peptide-coated gold particles were supplied to laboratories in two forms: a colloidal suspension in pure water and; particles dried onto a silicon wafer. Participants prepared and analyzed these samples using either X-ray photoelectron spectroscopy (XPS) or low energy ion scattering (LEIS). Careful data analysis revealed some significant sources of discrepancy, particularly for XPS. Degradation during transportation, storage or sample preparation resulted in a variability in thickness of 53 %. The calculation method chosen by XPS participants contributed a variability of 67 %. However, variability of 12 % was achieved for the samples deposited using a single method and by choosing photoelectron peaks that were not adversely affected by instrumental transmission effects. The study identified a need for more consistency in instrumental transmission functions and relative sensitivity factors, since this contributed a variability of 33 %. The results from the LEIS participants were more consistent, with variability of less than 10 % in thickness and this is mostly due to a common method of data analysis. The calculation was performed using a model developed for uniform, flat films and some participants employed a correction factor to account for the sample geometry, which appears warranted based upon a simulation of LEIS data from one of the participants and comparison to the XPS results.

Interlaboratory study for the validation of an ecotoxicological procedure to monitor the quality of septic sludge received at a wastewater treatment plant.

PubMed

Robidoux, P Y; Choucri, A; Bastien, C; Sunahara, G I; López-Gastey, J

2001-01-01

Septic tank sludge is regularly hauled to the Montreal Urban Community (MUC) wastewater treatment plant. It is then discharged and mixed with the wastewater inflow before entering the primary chemical treatment process. An ecotoxicological procedure integrating chemical and toxicological analyses has been recently developed and applied to screen for the illicit discharge of toxic substances in septic sludge. The toxicity tests used were the Microtox, the bacterial-respiration, and the lettuce (Lactuca sativa) root elongation tests. In order to validate the applicability of the proposed procedure, a two-year interlaboratory study was carried out. In general, the results obtained by two independent laboratories (MUC and the Centre d'expertise en analyse environnementale du Quebec) were comparable and reproducible. Some differences were found using the Microtox test. Organic (e.g., phenol and formaldehyde) and inorganic (e.g., nickel and cyanide) spiked septic sludge were detected with good reliability and high efficiency. The relative efficiency to detect spiked substances was > 70% and confirms the results of previous studies. In addition, the respiration test was the most efficient toxicological tool to detect spiked substances, whereas the Microtox was the least efficient (< 15%). Efficiencies to detect spiked contaminants were also similar for both laboratories. These results support previous data presented earlier and contribute to the validation of the ecotoxicological procedure used by the MUC to screen toxicity in septic sludge.

Genome-scale cluster analysis of replicated microarrays using shrinkage correlation coefficient.

PubMed

Yao, Jianchao; Chang, Chunqi; Salmi, Mari L; Hung, Yeung Sam; Loraine, Ann; Roux, Stanley J

2008-06-18

Currently, clustering with some form of correlation coefficient as the gene similarity metric has become a popular method for profiling genomic data. The Pearson correlation coefficient and the standard deviation (SD)-weighted correlation coefficient are the two most widely-used correlations as the similarity metrics in clustering microarray data. However, these two correlations are not optimal for analyzing replicated microarray data generated by most laboratories. An effective correlation coefficient is needed to provide statistically sufficient analysis of replicated microarray data. In this study, we describe a novel correlation coefficient, shrinkage correlation coefficient (SCC), that fully exploits the similarity between the replicated microarray experimental samples. The methodology considers both the number of replicates and the variance within each experimental group in clustering expression data, and provides a robust statistical estimation of the error of replicated microarray data. The value of SCC is revealed by its comparison with two other correlation coefficients that are currently the most widely-used (Pearson correlation coefficient and SD-weighted correlation coefficient) using statistical measures on both synthetic expression data as well as real gene expression data from Saccharomyces cerevisiae. Two leading clustering methods, hierarchical and k-means clustering were applied for the comparison. The comparison indicated that using SCC achieves better clustering performance. Applying SCC-based hierarchical clustering to the replicated microarray data obtained from germinating spores of the fern Ceratopteris richardii, we discovered two clusters of genes with shared expression patterns during spore germination. Functional analysis suggested that some of the genetic mechanisms that control germination in such diverse plant lineages as mosses and angiosperms are also conserved among ferns. This study shows that SCC is an alternative to the Pearson correlation coefficient and the SD-weighted correlation coefficient, and is particularly useful for clustering replicated microarray data. This computational approach should be generally useful for proteomic data or other high-throughput analysis methodology.

Comprehensive analysis of correlation coefficients estimated from pooling heterogeneous microarray data

PubMed Central

2013-01-01

Background The synthesis of information across microarray studies has been performed by combining statistical results of individual studies (as in a mosaic), or by combining data from multiple studies into a large pool to be analyzed as a single data set (as in a melting pot of data). Specific issues relating to data heterogeneity across microarray studies, such as differences within and between labs or differences among experimental conditions, could lead to equivocal results in a melting pot approach. Results We applied statistical theory to determine the specific effect of different means and heteroskedasticity across 19 groups of microarray data on the sign and magnitude of gene-to-gene Pearson correlation coefficients obtained from the pool of 19 groups. We quantified the biases of the pooled coefficients and compared them to the biases of correlations estimated by an effect-size model. Mean differences across the 19 groups were the main factor determining the magnitude and sign of the pooled coefficients, which showed largest values of bias as they approached ±1. Only heteroskedasticity across the pool of 19 groups resulted in less efficient estimations of correlations than did a classical meta-analysis approach of combining correlation coefficients. These results were corroborated by simulation studies involving either mean differences or heteroskedasticity across a pool of N > 2 groups. Conclusions The combination of statistical results is best suited for synthesizing the correlation between expression profiles of a gene pair across several microarray studies. PMID:23822712

Gene response profiles for Daphnia pulex exposed to the environmental stressor cadmium reveals novel crustacean metallothioneins.

PubMed

Shaw, Joseph R; Colbourne, John K; Davey, Jennifer C; Glaholt, Stephen P; Hampton, Thomas H; Chen, Celia Y; Folt, Carol L; Hamilton, Joshua W

2007-12-21

Genomic research tools such as microarrays are proving to be important resources to study the complex regulation of genes that respond to environmental perturbations. A first generation cDNA microarray was developed for the environmental indicator species Daphnia pulex, to identify genes whose regulation is modulated following exposure to the metal stressor cadmium. Our experiments revealed interesting changes in gene transcription that suggest their biological roles and their potentially toxicological features in responding to this important environmental contaminant. Our microarray identified genes reported in the literature to be regulated in response to cadmium exposure, suggested functional attributes for genes that share no sequence similarity to proteins in the public databases, and pointed to genes that are likely members of expanded gene families in the Daphnia genome. Genes identified on the microarray also were associated with cadmium induced phenotypes and population-level outcomes that we experimentally determined. A subset of genes regulated in response to cadmium exposure was independently validated using quantitative-realtime (Q-RT)-PCR. These microarray studies led to the discovery of three genes coding for the metal detoxication protein metallothionein (MT). The gene structures and predicted translated sequences of D. pulex MTs clearly place them in this gene family. Yet, they share little homology with previously characterized MTs. The genomic information obtained from this study represents an important first step in characterizing microarray patterns that may be diagnostic to specific environmental contaminants and give insights into their toxicological mechanisms, while also providing a practical tool for evolutionary, ecological, and toxicological functional gene discovery studies. Advances in Daphnia genomics will enable the further development of this species as a model organism for the environmental sciences.

Gene response profiles for Daphnia pulex exposed to the environmental stressor cadmium reveals novel crustacean metallothioneins

PubMed Central

Shaw, Joseph R; Colbourne, John K; Davey, Jennifer C; Glaholt, Stephen P; Hampton, Thomas H; Chen, Celia Y; Folt, Carol L; Hamilton, Joshua W

2007-01-01

Background Genomic research tools such as microarrays are proving to be important resources to study the complex regulation of genes that respond to environmental perturbations. A first generation cDNA microarray was developed for the environmental indicator species Daphnia pulex, to identify genes whose regulation is modulated following exposure to the metal stressor cadmium. Our experiments revealed interesting changes in gene transcription that suggest their biological roles and their potentially toxicological features in responding to this important environmental contaminant. Results Our microarray identified genes reported in the literature to be regulated in response to cadmium exposure, suggested functional attributes for genes that share no sequence similarity to proteins in the public databases, and pointed to genes that are likely members of expanded gene families in the Daphnia genome. Genes identified on the microarray also were associated with cadmium induced phenotypes and population-level outcomes that we experimentally determined. A subset of genes regulated in response to cadmium exposure was independently validated using quantitative-realtime (Q-RT)-PCR. These microarray studies led to the discovery of three genes coding for the metal detoxication protein metallothionein (MT). The gene structures and predicted translated sequences of D. pulex MTs clearly place them in this gene family. Yet, they share little homology with previously characterized MTs. Conclusion The genomic information obtained from this study represents an important first step in characterizing microarray patterns that may be diagnostic to specific environmental contaminants and give insights into their toxicological mechanisms, while also providing a practical tool for evolutionary, ecological, and toxicological functional gene discovery studies. Advances in Daphnia genomics will enable the further development of this species as a model organism for the environmental sciences. PMID:18154678

Inter-laboratory study to characterize the detection of serum antibodies against porcine epidemic diarrhoea virus.

PubMed

Strandbygaard, Bertel; Lavazza, Antonio; Lelli, Davide; Blanchard, Yannick; Grasland, Béatrice; Poder, Sophie Le; Rose, Nicolas; Steinbach, Falko; van der Poel, Wim H M; Widén, Frederik; Belsham, Graham J; Bøtner, Anette

2016-12-25

Porcine epidemic diarrhea virus (PEDV) has caused extensive economic losses to pig producers in many countries. It was recently introduced, for the first time, into North America and outbreaks have occurred again in multiple countries within Europe as well. To assess the properties of various diagnostic assays for the detection of PEDV infection, multiple panels of porcine sera have been shared and tested for the presence of antibodies against PEDV in an inter-laboratory ring trial. Different laboratories have used a variety of "in house" ELISAs and also one commercial assay. The sensitivity and specificity of each assay has been estimated using a Bayesian analysis applied to the ring trial results obtained with the different assays in the absence of a gold standard. Although different characteristics were found, it can be concluded that each of the assays used can detect infection of pigs at a herd level by either the early European strains of PEDV or the recently circulating strains (INDEL and non-INDEL). However, not all the assays seem suitable for demonstrating freedom from disease in a country. The results from individual animals, especially when the infection has occurred within an experimental situation, show more variation. Copyright © 2016. Published by Elsevier B.V.

Commutability of NIST SRM 1955 Homocysteine and Folate in Frozen Human Serum with selected total homocysteine immunoassays and enzymatic assays.

PubMed

Nelson, Bryant C; Pfeiffer, Christine M; Zhang, Ming; Duewer, David L; Sharpless, Katherine E; Lippa, Katrice A

2008-09-01

The National Institute of Standards and Technology (NIST) has recently developed Standard Reference Material (SRM) 1955 Homocysteine and Folate in Frozen Human Serum with certified values for total homocysteine (tHcy) and 5-methyl-tetrahydrofolic acid. NIST has performed an international, interlaboratory assessment of SRM 1955 commutability; results are reported for tHcy only. Total Hcy was measured in 20 patient sera and in 3 levels of SRM 1955 using 14 immunoassays and/or enzymatic assays. Liquid chromatography/tandem mass spectrometry was utilized as the reference assay. An "errors-in-variables" statistical model was utilized to assess the commutability of SRM 1955. Normalized residuals ranged from -2.65 to 2.19 for SRM 1955. The median interlaboratory/interassay imprecision (CV) was approximately 4% for patient specimens and ranged from approximately 3% to approximately 7% for SRM 1955. The median intra-assay imprecision ranged from approximately 1% to approximately 13%. Orthogonal residuals, as a descriptor of assay accuracy, ranged from 0.29 to 7.71 and from 0.20 to 2.22 for patient specimens and SRM 1955 samples, respectively. The current study suggests that SRM 1955 is commutable with the investigated tHcy assays; however, a broader specimen set needs to be evaluated to completely substantiate this conclusion.

Construction of measurement uncertainty profiles for quantitative analysis of genetically modified organisms based on interlaboratory validation data.

PubMed

Macarthur, Roy; Feinberg, Max; Bertheau, Yves

2010-01-01

A method is presented for estimating the size of uncertainty associated with the measurement of products derived from genetically modified organisms (GMOs). The method is based on the uncertainty profile, which is an extension, for the estimation of uncertainty, of a recent graphical statistical tool called an accuracy profile that was developed for the validation of quantitative analytical methods. The application of uncertainty profiles as an aid to decision making and assessment of fitness for purpose is also presented. Results of the measurement of the quantity of GMOs in flour by PCR-based methods collected through a number of interlaboratory studies followed the log-normal distribution. Uncertainty profiles built using the results generally give an expected range for measurement results of 50-200% of reference concentrations for materials that contain at least 1% GMO. This range is consistent with European Network of GM Laboratories and the European Union (EU) Community Reference Laboratory validation criteria and can be used as a fitness for purpose criterion for measurement methods. The effect on the enforcement of EU labeling regulations is that, in general, an individual analytical result needs to be < 0.45% to demonstrate compliance, and > 1.8% to demonstrate noncompliance with a labeling threshold of 0.9%.

[Preparation of the cDNA microarray on the differential expressed cDNA of senescence-accelerated mouse's hippocampus].

PubMed

Cheng, Xiao-Rui; Zhou, Wen-Xia; Zhang, Yong-Xiang

2006-05-01

Alzheimer' s disease (AD) is the most common form of dementia in the elderly. AD is an invariably fatal neurodegenerative disorder with no effective treatment. Senescence-accelerated mouse prone 8 (SAMP8) is a model for studying age-related cognitive impairments and also is a good model to study brain aging and one of mouse model of AD. The technique of cDNA microarray can monitor the expression levels of thousands of genes simultaneously and can be used to study AD with the character of multi-mechanism, multi-targets and multi-pathway. In order to disclose the mechanism of AD and find the drug targets of AD, cDNA microarray containing 3136 cDNAs amplified from the suppression subtracted cDNA library of hippocampus of SAMP8 and SAMR1 was prepared with 16 blocks and 14 x 14 pins, the housekeeping gene beta-actin and G3PDH as inner conference. The background of this microarray was low and unanimous, and dots divided evenly. The conditions of hybridization and washing were optimized during the hybridization of probe and target molecule. After the data of hybridization analysis, the differential expressed cDNAs were sequenced and analyzed by the bioinformatics, and some of genes were quantified by the real time RT-PCR and the reliability of this cDNA microarray were validated. This cDNA microarray may be the good means to select the differential expressed genes and disclose the molecular mechanism of SAMP8's brain aging and AD.

DNA Microarray Detection of 18 Important Human Blood Protozoan Species

PubMed Central

Chen, Jun-Hu; Feng, Xin-Yu; Chen, Shao-Hong; Cai, Yu-Chun; Lu, Yan; Zhou, Xiao-Nong; Chen, Jia-Xu; Hu, Wei

2016-01-01

Background Accurate detection of blood protozoa from clinical samples is important for diagnosis, treatment and control of related diseases. In this preliminary study, a novel DNA microarray system was assessed for the detection of Plasmodium, Leishmania, Trypanosoma, Toxoplasma gondii and Babesia in humans, animals, and vectors, in comparison with microscopy and PCR data. Developing a rapid, simple, and convenient detection method for protozoan detection is an urgent need. Methodology/Principal Findings The microarray assay simultaneously identified 18 species of common blood protozoa based on the differences in respective target genes. A total of 20 specific primer pairs and 107 microarray probes were selected according to conserved regions which were designed to identify 18 species in 5 blood protozoan genera. The positive detection rate of the microarray assay was 91.78% (402/438). Sensitivity and specificity for blood protozoan detection ranged from 82.4% (95%CI: 65.9% ~ 98.8%) to 100.0% and 95.1% (95%CI: 93.2% ~ 97.0%) to 100.0%, respectively. Positive predictive value (PPV) and negative predictive value (NPV) ranged from 20.0% (95%CI: 2.5% ~ 37.5%) to 100.0% and 96.8% (95%CI: 95.0% ~ 98.6%) to 100.0%, respectively. Youden index varied from 0.82 to 0.98. The detection limit of the DNA microarrays ranged from 200 to 500 copies/reaction, similar to PCR findings. The concordance rate between microarray data and DNA sequencing results was 100%. Conclusions/Significance Overall, the newly developed microarray platform provides a convenient, highly accurate, and reliable clinical assay for the determination of blood protozoan species. PMID:27911895

The Diagnostic Yield of Array Comparative Genomic Hybridization Is High Regardless of Severity of Intellectual Disability/Developmental Delay in Children.

PubMed

D'Arrigo, Stefano; Gavazzi, Francesco; Alfei, Enrico; Zuffardi, Orsetta; Montomoli, Cristina; Corso, Barbara; Buzzi, Erika; Sciacca, Francesca L; Bulgheroni, Sara; Riva, Daria; Pantaleoni, Chiara

2016-05-01

Microarray-based comparative genomic hybridization is a method of molecular analysis that identifies chromosomal anomalies (or copy number variants) that correlate with clinical phenotypes. The aim of the present study was to apply a clinical score previously designated by de Vries to 329 patients with intellectual disability/developmental disorder (intellectual disability/developmental delay) referred to our tertiary center and to see whether the clinical factors are associated with a positive outcome of aCGH analyses. Another goal was to test the association between a positive microarray-based comparative genomic hybridization result and the severity of intellectual disability/developmental delay. Microarray-based comparative genomic hybridization identified structural chromosomal alterations responsible for the intellectual disability/developmental delay phenotype in 16% of our sample. Our study showed that causative copy number variants are frequently found even in cases of mild intellectual disability (30.77%). We want to emphasize the need to conduct microarray-based comparative genomic hybridization on all individuals with intellectual disability/developmental delay, regardless of the severity, because the degree of intellectual disability/developmental delay does not predict the diagnostic yield of microarray-based comparative genomic hybridization. © The Author(s) 2015.

Recognition of multiple imbalanced cancer types based on DNA microarray data using ensemble classifiers.

PubMed

Yu, Hualong; Hong, Shufang; Yang, Xibei; Ni, Jun; Dan, Yuanyuan; Qin, Bin

2013-01-01

DNA microarray technology can measure the activities of tens of thousands of genes simultaneously, which provides an efficient way to diagnose cancer at the molecular level. Although this strategy has attracted significant research attention, most studies neglect an important problem, namely, that most DNA microarray datasets are skewed, which causes traditional learning algorithms to produce inaccurate results. Some studies have considered this problem, yet they merely focus on binary-class problem. In this paper, we dealt with multiclass imbalanced classification problem, as encountered in cancer DNA microarray, by using ensemble learning. We utilized one-against-all coding strategy to transform multiclass to multiple binary classes, each of them carrying out feature subspace, which is an evolving version of random subspace that generates multiple diverse training subsets. Next, we introduced one of two different correction technologies, namely, decision threshold adjustment or random undersampling, into each training subset to alleviate the damage of class imbalance. Specifically, support vector machine was used as base classifier, and a novel voting rule called counter voting was presented for making a final decision. Experimental results on eight skewed multiclass cancer microarray datasets indicate that unlike many traditional classification approaches, our methods are insensitive to class imbalance.

Comprehensive Assessments of RNA-seq by the SEQC Consortium: FDA-Led Efforts Advance Precision Medicine.

PubMed

Xu, Joshua; Gong, Binsheng; Wu, Leihong; Thakkar, Shraddha; Hong, Huixiao; Tong, Weida

2016-03-15

Studies on gene expression in response to therapy have led to the discovery of pharmacogenomics biomarkers and advances in precision medicine. Whole transcriptome sequencing (RNA-seq) is an emerging tool for profiling gene expression and has received wide adoption in the biomedical research community. However, its value in regulatory decision making requires rigorous assessment and consensus between various stakeholders, including the research community, regulatory agencies, and industry. The FDA-led SEquencing Quality Control (SEQC) consortium has made considerable progress in this direction, and is the subject of this review. Specifically, three RNA-seq platforms (Illumina HiSeq, Life Technologies SOLiD, and Roche 454) were extensively evaluated at multiple sites to assess cross-site and cross-platform reproducibility. The results demonstrated that relative gene expression measurements were consistently comparable across labs and platforms, but not so for the measurement of absolute expression levels. As part of the quality evaluation several studies were included to evaluate the utility of RNA-seq in clinical settings and safety assessment. The neuroblastoma study profiled tumor samples from 498 pediatric neuroblastoma patients by both microarray and RNA-seq. RNA-seq offers more utilities than microarray in determining the transcriptomic characteristics of cancer. However, RNA-seq and microarray-based models were comparable in clinical endpoint prediction, even when including additional features unique to RNA-seq beyond gene expression. The toxicogenomics study compared microarray and RNA-seq profiles of the liver samples from rats exposed to 27 different chemicals representing multiple toxicity modes of action. Cross-platform concordance was dependent on chemical treatment and transcript abundance. Though both RNA-seq and microarray are suitable for developing gene expression based predictive models with comparable prediction performance, RNA-seq offers advantages over microarray in profiling genes with low expression. The rat BodyMap study provided a comprehensive rat transcriptomic body map by performing RNA-Seq on 320 samples from 11 organs in either sex of juvenile, adolescent, adult and aged Fischer 344 rats. Lastly, the transferability study demonstrated that signature genes of predictive models are reciprocally transferable between microarray and RNA-seq data for model development using a comprehensive approach with two large clinical data sets. This result suggests continued usefulness of legacy microarray data in the coming RNA-seq era. In conclusion, the SEQC project enhances our understanding of RNA-seq and provides valuable guidelines for RNA-seq based clinical application and safety evaluation to advance precision medicine.

16S rRNA gene-based phylogenetic microarray for simultaneous identification of members of the genus Burkholderia.

PubMed

Schönmann, Susan; Loy, Alexander; Wimmersberger, Céline; Sobek, Jens; Aquino, Catharine; Vandamme, Peter; Frey, Beat; Rehrauer, Hubert; Eberl, Leo

2009-04-01

For cultivation-independent and highly parallel analysis of members of the genus Burkholderia, an oligonucleotide microarray (phylochip) consisting of 131 hierarchically nested 16S rRNA gene-targeted oligonucleotide probes was developed. A novel primer pair was designed for selective amplification of a 1.3 kb 16S rRNA gene fragment of Burkholderia species prior to microarray analysis. The diagnostic performance of the microarray for identification and differentiation of Burkholderia species was tested with 44 reference strains of the genera Burkholderia, Pandoraea, Ralstonia and Limnobacter. Hybridization patterns based on presence/absence of probe signals were interpreted semi-automatically using the novel likelihood-based strategy of the web-tool Phylo- Detect. Eighty-eight per cent of the reference strains were correctly identified at the species level. The evaluated microarray was applied to investigate shifts in the Burkholderia community structure in acidic forest soil upon addition of cadmium, a condition that selected for Burkholderia species. The microarray results were in agreement with those obtained from phylogenetic analysis of Burkholderia 16S rRNA gene sequences recovered from the same cadmiumcontaminated soil, demonstrating the value of the Burkholderia phylochip for determinative and environmental studies.

Determination of Minimum Training Sample Size for Microarray-Based Cancer Outcome Prediction–An Empirical Assessment

PubMed Central

Cheng, Ningtao; Wu, Leihong; Cheng, Yiyu

2013-01-01

The promise of microarray technology in providing prediction classifiers for cancer outcome estimation has been confirmed by a number of demonstrable successes. However, the reliability of prediction results relies heavily on the accuracy of statistical parameters involved in classifiers. It cannot be reliably estimated with only a small number of training samples. Therefore, it is of vital importance to determine the minimum number of training samples and to ensure the clinical value of microarrays in cancer outcome prediction. We evaluated the impact of training sample size on model performance extensively based on 3 large-scale cancer microarray datasets provided by the second phase of MicroArray Quality Control project (MAQC-II). An SSNR-based (scale of signal-to-noise ratio) protocol was proposed in this study for minimum training sample size determination. External validation results based on another 3 cancer datasets confirmed that the SSNR-based approach could not only determine the minimum number of training samples efficiently, but also provide a valuable strategy for estimating the underlying performance of classifiers in advance. Once translated into clinical routine applications, the SSNR-based protocol would provide great convenience in microarray-based cancer outcome prediction in improving classifier reliability. PMID:23861920

2008 Microarray Research Group (MARG Survey): Sensing the State of Microarray Technology

EPA Science Inventory

Over the past several years, the field of microarrays has grown and evolved drastically. In its continued efforts to track this evolution and transformation, the ABRF-MARG has once again conducted a survey of international microarray facilities and individual microarray users. Th...

Performance of automated scoring of ER, PR, HER2, CK5/6 and EGFR in breast cancer tissue microarrays in the Breast Cancer Association Consortium

PubMed Central

Howat, William J; Blows, Fiona M; Provenzano, Elena; Brook, Mark N; Morris, Lorna; Gazinska, Patrycja; Johnson, Nicola; McDuffus, Leigh‐Anne; Miller, Jodi; Sawyer, Elinor J; Pinder, Sarah; van Deurzen, Carolien H M; Jones, Louise; Sironen, Reijo; Visscher, Daniel; Caldas, Carlos; Daley, Frances; Coulson, Penny; Broeks, Annegien; Sanders, Joyce; Wesseling, Jelle; Nevanlinna, Heli; Fagerholm, Rainer; Blomqvist, Carl; Heikkilä, Päivi; Ali, H Raza; Dawson, Sarah‐Jane; Figueroa, Jonine; Lissowska, Jolanta; Brinton, Louise; Mannermaa, Arto; Kataja, Vesa; Kosma, Veli‐Matti; Cox, Angela; Brock, Ian W; Cross, Simon S; Reed, Malcolm W; Couch, Fergus J; Olson, Janet E; Devillee, Peter; Mesker, Wilma E; Seyaneve, Caroline M; Hollestelle, Antoinette; Benitez, Javier; Perez, Jose Ignacio Arias; Menéndez, Primitiva; Bolla, Manjeet K; Easton, Douglas F; Schmidt, Marjanka K; Pharoah, Paul D; Sherman, Mark E

2014-01-01

Abstract Breast cancer risk factors and clinical outcomes vary by tumour marker expression. However, individual studies often lack the power required to assess these relationships, and large‐scale analyses are limited by the need for high throughput, standardized scoring methods. To address these limitations, we assessed whether automated image analysis of immunohistochemically stained tissue microarrays can permit rapid, standardized scoring of tumour markers from multiple studies. Tissue microarray sections prepared in nine studies containing 20 263 cores from 8267 breast cancers stained for two nuclear (oestrogen receptor, progesterone receptor), two membranous (human epidermal growth factor receptor 2 and epidermal growth factor receptor) and one cytoplasmic (cytokeratin 5/6) marker were scanned as digital images. Automated algorithms were used to score markers in tumour cells using the Ariol system. We compared automated scores against visual reads, and their associations with breast cancer survival. Approximately 65–70% of tissue microarray cores were satisfactory for scoring. Among satisfactory cores, agreement between dichotomous automated and visual scores was highest for oestrogen receptor (Kappa = 0.76), followed by human epidermal growth factor receptor 2 (Kappa = 0.69) and progesterone receptor (Kappa = 0.67). Automated quantitative scores for these markers were associated with hazard ratios for breast cancer mortality in a dose‐response manner. Considering visual scores of epidermal growth factor receptor or cytokeratin 5/6 as the reference, automated scoring achieved excellent negative predictive value (96–98%), but yielded many false positives (positive predictive value = 30–32%). For all markers, we observed substantial heterogeneity in automated scoring performance across tissue microarrays. Automated analysis is a potentially useful tool for large‐scale, quantitative scoring of immunohistochemically stained tissue microarrays available in consortia. However, continued optimization, rigorous marker‐specific quality control measures and standardization of tissue microarray designs, staining and scoring protocols is needed to enhance results. PMID:27499890

Round-robin 230Th– 234U age dating of bulk uranium for nuclear forensics

DOE PAGES

Gaffney, Amy M.; Hubert, Amélie; Kinman, William S.; ...

2015-07-30

We report that in an inter-laboratory measurement comparison study, four laboratories determined 230Th– 234U model ages of uranium certified reference material NBL U050 using isotope dilution mass spectrometry. The model dates determined by the participating laboratories range from 9 March 1956 to 19 October 1957, and are indistinguishable given the associated measurement uncertainties. These model ages are concordant with to slightly older than the known production age of NBL U050.

Round-robin 230Th– 234U age dating of bulk uranium for nuclear forensics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gaffney, Amy M.; Hubert, Amélie; Kinman, William S.

We report that in an inter-laboratory measurement comparison study, four laboratories determined 230Th– 234U model ages of uranium certified reference material NBL U050 using isotope dilution mass spectrometry. The model dates determined by the participating laboratories range from 9 March 1956 to 19 October 1957, and are indistinguishable given the associated measurement uncertainties. These model ages are concordant with to slightly older than the known production age of NBL U050.

Interlaboratory Comparison of Quantitative PCR Test Results for Dehalococcoides

EPA Science Inventory

Quantitative PCR (qPCR) techniques have been widely used to measure Dehalococcoides (Dhc) DNA in the groundwater at field sites for several years. Interpretation of these data may be complicated when different laboratories using alternate methods conduct the analysis. An...

Tools to minimize interlaboratory variability in vitellogenin gene expression monitoring programs

EPA Science Inventory

The egg yolk precursor protein vitellogenin is widely used as a biomarker of estrogen exposure in male fish. However, standardized methodology is lacking and little is known regarding the reproducibility of results among laboratories using different equipment, reagents, protocols...

Interlaboratory Variability of Slip Coefficient Testing for Bridge Coatings

DOT National Transportation Integrated Search

2014-12-01

All steel bridge systems need some type of a corrosion protection scheme to ensure a serviceable life. The most common approach is to use a multilayered paint system with a zinc-rich primer. In addition to corrosion performance, other factors need to...

THE ABRF-MARG MICROARRAY SURVEY 2004: TAKING THE PULSE OF THE MICROARRAY FIELD

EPA Science Inventory

Over the past several years, the field of microarrays has grown and evolved drastically. In its continued efforts to track this evolution, the ABRF-MARG has once again conducted a survey of international microarray facilities and individual microarray users. The goal of the surve...

Parallel, confocal, and complete spectrum imager for fluorescent detection of high-density microarray

NASA Astrophysics Data System (ADS)

Bogdanov, Valery L.; Boyce-Jacino, Michael

1999-05-01

Confined arrays of biochemical probes deposited on a solid support surface (analytical microarray or 'chip') provide an opportunity to analysis multiple reactions simultaneously. Microarrays are increasingly used in genetics, medicine and environment scanning as research and analytical instruments. A power of microarray technology comes from its parallelism which grows with array miniaturization, minimization of reagent volume per reaction site and reaction multiplexing. An optical detector of microarray signals should combine high sensitivity, spatial and spectral resolution. Additionally, low-cost and a high processing rate are needed to transfer microarray technology into biomedical practice. We designed an imager that provides confocal and complete spectrum detection of entire fluorescently-labeled microarray in parallel. Imager uses microlens array, non-slit spectral decomposer, and high- sensitive detector (cooled CCD). Two imaging channels provide a simultaneous detection of localization, integrated and spectral intensities for each reaction site in microarray. A dimensional matching between microarray and imager's optics eliminates all in moving parts in instrumentation, enabling highly informative, fast and low-cost microarray detection. We report theory of confocal hyperspectral imaging with microlenses array and experimental data for implementation of developed imager to detect fluorescently labeled microarray with a density approximately 103 sites per cm2.

Generalized Correlation Coefficient for Non-Parametric Analysis of Microarray Time-Course Data.

PubMed

Tan, Qihua; Thomassen, Mads; Burton, Mark; Mose, Kristian Fredløv; Andersen, Klaus Ejner; Hjelmborg, Jacob; Kruse, Torben

2017-06-06

Modeling complex time-course patterns is a challenging issue in microarray study due to complex gene expression patterns in response to the time-course experiment. We introduce the generalized correlation coefficient and propose a combinatory approach for detecting, testing and clustering the heterogeneous time-course gene expression patterns. Application of the method identified nonlinear time-course patterns in high agreement with parametric analysis. We conclude that the non-parametric nature in the generalized correlation analysis could be an useful and efficient tool for analyzing microarray time-course data and for exploring the complex relationships in the omics data for studying their association with disease and health.

Development of the first oligonucleotide microarray for global gene expression profiling in guinea pigs: defining the transcription signature of infectious diseases.

PubMed

Jain, Ruchi; Dey, Bappaditya; Tyagi, Anil K

2012-10-02

The Guinea pig (Cavia porcellus) is one of the most extensively used animal models to study infectious diseases. However, despite its tremendous contribution towards understanding the establishment, progression and control of a number of diseases in general and tuberculosis in particular, the lack of fully annotated guinea pig genome sequence as well as appropriate molecular reagents has severely hampered detailed genetic and immunological analysis in this animal model. By employing the cross-species hybridization technique, we have developed an oligonucleotide microarray with 44,000 features assembled from different mammalian species, which to the best of our knowledge is the first attempt to employ microarray to study the global gene expression profile in guinea pigs. To validate and demonstrate the merit of this microarray, we have studied, as an example, the expression profile of guinea pig lungs during the advanced phase of M. tuberculosis infection. A significant upregulation of 1344 genes and a marked down regulation of 1856 genes in the lungs identified a disease signature of pulmonary tuberculosis infection. We report the development of first comprehensive microarray for studying the global gene expression profile in guinea pigs and validation of its usefulness with tuberculosis as a case study. An important gap in the area of infectious diseases has been addressed and a valuable molecular tool is provided to optimally harness the potential of guinea pig model to develop better vaccines and therapies against human diseases.

CONFIRMING MICROARRAY DATA--IS IT REALLY NECESSARY?

EPA Science Inventory

The generation of corroborative data has become a commonly used approach for ensuring the veracity of microarray data. Indeed, the need to conduct corroborative studies has now become official editorial policy for at least two journals, and several more are considering introducin...

Interlaboratory variability in the quantification of new generation antiepileptic drugs based on external quality assessment data.

PubMed

Williams, John; Bialer, Meir; Johannessen, Svein I; Krämer, Günther; Levy, René; Mattson, Richard H; Perucca, Emilio; Patsalos, Philip N; Wilson, John F

2003-01-01

To assess interlaboratory variability in the determination of serum levels of new antiepileptic drugs (AEDs). Lyophilised serum samples containing clinically relevant concentrations of felbamate (FBM), gabapentin (GBP), lamotrigine (LTG), the monohydroxy derivative of oxcarbazepine (OCBZ; MHD), tiagabine (TGB), topiramate (TPM), and vigabatrin (VGB) were distributed monthly among 70 laboratories participating in the international Heathcontrol External Quality Assessment Scheme (EQAS). Assay results returned over a 15-month period were evaluated for precision and accuracy. The most frequently measured compound was LTG (65), followed by MHD (39), GBP (19), TPM (18), VGB (15), FBM (16), and TGB (8). High-performance liquid chromatography was the most commonly used assay technique for all drugs except for TPM, for which two thirds of laboratories used a commercial immunoassay. For all assay methods combined, precision was <11% for MHD, FBM, TPM, and LTG, close to 15% for GBP and VGB, and as high as 54% for TGB (p < 0.001). Mean accuracy values were <10% for all drugs other than TGB, for which measured values were on average 13.9% higher than spiked values, with a high variability around the mean (45%). No differences in precision and accuracy were found between methods, except for TPM, for which gas chromatography showed poorer accuracy compared with immunoassay and gas chromatography-mass spectrometry. With the notable exception of TGB, interlaboratory variability in the determination of new AEDs was comparable to that reported with older-generation agents. Poor assay performance is related more to individual operators than to the intrinsic characteristics of the method applied. Participation in an EQAS scheme is recommended to ensure adequate control of assay variability in therapeutic drug monitoring.

First European interlaboratory comparison of tetracycline and age determination with red fox teeth following oral rabies vaccination programs.

PubMed

Robardet, Emmanuelle; Demerson, Jean-Michel; Andrieu, Sabrina; Cliquet, Florence

2012-10-01

The first European interlaboratory comparison of tetracycline and age determination with red fox (Vulpes vulpes) tooth samples was organized by the European Union Reference Laboratory for rabies. Performance and procedures implemented by member states were compared. These techniques are widely used to monitor bait uptake in European oral rabies vaccination campaigns. A panel of five red fox half-mandibles comprising one weak positive juvenile sample, two positive adult samples, one negative juvenile sample, and one negative adult sample were sent, along with a technical questionnaire, to 12 laboratories participating on a voluntary basis. The results of only three laboratories (25%) were 100% correct. False-negative results were more frequently seen in weak positive juvenile samples (58%) but were infrequent in positive adult samples (4%), probably due to differences in the ease of reading the two groups of teeth. Four laboratories (44%) had correct results for age determination on all samples. Ages were incorrectly identified in both adult and juvenile samples, with 11 and 17% of discordant results, respectively. Analysis of the technical questionnaires in parallel with test results suggested that all laboratories cutting mandible sections between the canine and first premolar obtained false results. All the laboratories using longitudinal rather than transverse sections and those not using a mounting medium also produced false results. Section thickness appeared to affect the results; no mistakes were found in laboratories using sections <150 μm thick. Factors having a potential impact on the success of laboratories were discussed, and recommendations proposed. Such interlaboratory trials underline the importance of using standardized procedures for biomarker detection in oral rabies vaccination campaigns. Several changes can be made to improve analysis quality and increase the comparability of bait uptake frequencies among member states.

Extra-analytical quality indicators and laboratory performances.

PubMed

Sciacovelli, Laura; Aita, Ada; Plebani, Mario

2017-07-01

In the last few years much progress has been made in raising the awareness of laboratory medicine professionals about the effectiveness of quality indicators (QIs) in monitoring, and improving upon, performances in the extra-analytical phases of the Total Testing Process (TTP). An effective system for management of QIs includes the implementation of an internal assessment system and participation in inter-laboratory comparison. A well-designed internal assessment system allows the identification of critical activities and their systematic monitoring. Active participation in inter-laboratory comparison provides information on the performance level of one laboratory with respect to that of other participating laboratories. In order to guarantee the use of appropriate QIs and facilitate their implementation, many laboratories have adopted the Model of Quality Indicators (MQI) proposed by Working Group "Laboratory Errors and Patient Safety" (WG-LEPS) of IFCC, since 2008, which is the result of international consensus and continuous experimentation, and updating to meet new, constantly emerging needs. Data from participating laboratories are collected monthly and reports describing the statistical results and evaluating laboratory data, utilizing the Six Sigma metric, issued regularly. Although the results demonstrate that the processes need to be improved upon, overall the comparison with data collected in 2014 shows a general stability of quality levels and that an improvement has been achieved over time for some activities. The continuous monitoring of QI data allows identification all possible improvements, thus highlighting the value of participation in the inter-laboratory program proposed by WG-LEPS. The active participation of numerous laboratories will guarantee an ever more significant State-of-the-Art, promote the reduction of errors and improve quality of the TTP, thus guaranteeing patient safety. Copyright © 2017. Published by Elsevier Inc.

Quality of pathology reporting is crucial for cancer care and registration: a baseline assessment for breast cancers diagnosed in Belgium in 2008.

PubMed

De Schutter, H; Van Damme, N; Colpaert, C; Galant, C; Lambein, K; Cornelis, A; Neven, P; Van Eycken, E

2015-04-01

Given the crucial role of pathology reporting in the management of breast cancers, we aimed to investigate the quality and variability of breast cancer pathology reporting in Belgium. Detailed information on non-molecular and molecular parameters was retrieved from the pathology protocols available at the Belgian Cancer Registry for 10,007 breast cancers diagnosed in Belgium in 2008. Substantial underreporting was shown for several clinically relevant non-molecular parameters, such as lymphovascular invasion. High-volume laboratories performed only slightly better than others, and analyses at the individual laboratory level showed clear inter-laboratory variability in reporting for all volume categories. Information on ER/PR and HER2 IHC was mentioned in respectively 91.7% and 90.8% of evaluative cases. HER2 ISH data were available for 78.5% of the cases judged to be 2+ for HER2 IHC. For cases with different specimens analysed, discordance between these specimens was highest for HER2, followed by PR. For HER2, results obtained from different laboratories were even less concordant. In addition, inter-laboratory differences were noted in the used ER/PR scoring systems, the proportion of ER-/PR+ cases, and the relation between histological grade and ER/PR positivity. Data on Ki67 were only available for 43.8% of the investigated cases, and showed inconsistent use of cut-off values. Breast pathology reporting in Belgium in 2008 was suboptimal and showed considerable inter-laboratory variability. Synoptic reporting has been proposed as a facilitator towards increased reporting quality and harmonization, but the lack of aligned informatics remains a major hurdle in its concrete implementation. Copyright © 2014 Elsevier Ltd. All rights reserved.

Interlaboratory evaluation of Hyalella azteca and Chironomus tentans short-term and long-term sediment toxicity tests

USGS Publications Warehouse

Norberg-King, T. J.; Sibley, P.K.; Burton, G.A.; Ingersoll, C.G.; Kemble, N.E.; Ireland, S.; Mount, D.R.; Rowland, C.D.

2006-01-01

Methods for assessing the long-term toxicity of sediments to Hyalella azteca and Chironomus tentans can significantly enhance the capacity to assess sublethal effects of contaminated sediments through multiple endpoints. Sublethal tests allow us to begin to understand the relationship between short-term and long-term effects for toxic sediments. We present an interlaboratory evaluation with long-term and 10-d tests using control and contaminated sediments in which we assess whether proposed and existing performance criteria (test acceptability criteria [TAC]) could be achieved. Laboratories became familiar with newly developed, long-term protocols by testing two control sediments in phase 1. In phase 2, the 10-d and long-term tests were examined with several sediments. Laboratories met the TACs, but results varied depending on the test organism, test duration, and endpoints. For the long-term tests in phase 1, 66 to 100% of the laboratories consistently met the TACs for survival, growth, or reproduction using H. azteca, and 70 to 100% of the laboratories met the TACs for survival and growth, emergence, reproduction, and hatchability using C. tentans. In phase 2, fewer laboratories participated in long-term tests: 71 to 88% of the laboratories met the TAC for H. azteca, whereas 50 to 67% met the TAC for C. tentans. In the 10-d tests with H. azteca, and C. tentans, 82 and 88% of the laboratories met the TAC for survival, respectively, and 80% met the TAC for C. tentans growth. For the 10-d and long-term tests, laboratories predicted similar toxicity. Overall, the interlaboratory evaluation showed good precision of the methods, appropriate endpoints were incorporated into the test protocols, and tests effectively predicted the toxicity of sediments.

Fish and chips: Various methodologies demonstrate utility of a 16,006-gene salmonid microarray

PubMed Central

von Schalburg, Kristian R; Rise, Matthew L; Cooper, Glenn A; Brown, Gordon D; Gibbs, A Ross; Nelson, Colleen C; Davidson, William S; Koop, Ben F

2005-01-01

Background We have developed and fabricated a salmonid microarray containing cDNAs representing 16,006 genes. The genes spotted on the array have been stringently selected from Atlantic salmon and rainbow trout expressed sequence tag (EST) databases. The EST databases presently contain over 300,000 sequences from over 175 salmonid cDNA libraries derived from a wide variety of tissues and different developmental stages. In order to evaluate the utility of the microarray, a number of hybridization techniques and screening methods have been developed and tested. Results We have analyzed and evaluated the utility of a microarray containing 16,006 (16K) salmonid cDNAs in a variety of potential experimental settings. We quantified the amount of transcriptome binding that occurred in cross-species, organ complexity and intraspecific variation hybridization studies. We also developed a methodology to rapidly identify and confirm the contents of a bacterial artificial chromosome (BAC) library containing Atlantic salmon genomic DNA. Conclusion We validate and demonstrate the usefulness of the 16K microarray over a wide range of teleosts, even for transcriptome targets from species distantly related to salmonids. We show the potential of the use of the microarray in a variety of experimental settings through hybridization studies that examine the binding of targets derived from different organs and tissues. Intraspecific variation in transcriptome expression is evaluated and discussed. Finally, BAC hybridizations are demonstrated as a rapid and accurate means to identify gene content. PMID:16164747

Chemiluminescence microarrays in analytical chemistry: a critical review.

PubMed

Seidel, Michael; Niessner, Reinhard

2014-09-01

Multi-analyte immunoassays on microarrays and on multiplex DNA microarrays have been described for quantitative analysis of small organic molecules (e.g., antibiotics, drugs of abuse, small molecule toxins), proteins (e.g., antibodies or protein toxins), and microorganisms, viruses, and eukaryotic cells. In analytical chemistry, multi-analyte detection by use of analytical microarrays has become an innovative research topic because of the possibility of generating several sets of quantitative data for different analyte classes in a short time. Chemiluminescence (CL) microarrays are powerful tools for rapid multiplex analysis of complex matrices. A wide range of applications for CL microarrays is described in the literature dealing with analytical microarrays. The motivation for this review is to summarize the current state of CL-based analytical microarrays. Combining analysis of different compound classes on CL microarrays reduces analysis time, cost of reagents, and use of laboratory space. Applications are discussed, with examples from food safety, water safety, environmental monitoring, diagnostics, forensics, toxicology, and biosecurity. The potential and limitations of research on multiplex analysis by use of CL microarrays are discussed in this review.

Analysis of High-Throughput ELISA Microarray Data

DOE Office of Scientific and Technical Information (OSTI.GOV)

White, Amanda M.; Daly, Don S.; Zangar, Richard C.

Our research group develops analytical methods and software for the high-throughput analysis of quantitative enzyme-linked immunosorbent assay (ELISA) microarrays. ELISA microarrays differ from DNA microarrays in several fundamental aspects and most algorithms for analysis of DNA microarray data are not applicable to ELISA microarrays. In this review, we provide an overview of the steps involved in ELISA microarray data analysis and how the statistically sound algorithms we have developed provide an integrated software suite to address the needs of each data-processing step. The algorithms discussed are available in a set of open-source software tools (http://www.pnl.gov/statistics/ProMAT).

MICROARRAY ANALYSIS OF DICHLOROACETIC ACID-INDUCED CHANGES IN GENE EXPRESSION

EPA Science Inventory

MICROARRAY ANALYSIS OF DICHLOROACETIC ACID-INDUCED CHANGES IN GENE EXPRESSION

Dichloroacetic acid (DCA) is a major by-product of water disinfection by chlorination. Several studies have demonstrated the hepatocarcinogenicity of DCA in rodents when administered in dri...

Development of DNA Microarrays for Metabolic Pathway and Bioprocess Monitoring

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gregory Stephanopoulos

Transcriptional profiling experiments utilizing DNA microarrays to study the intracellular accumulation of PHB in Synechocystis has proved difficult in large part because strains that show significant differences in PHB which would justify global analysis of gene expression have not been isolated.

A meta-data based method for DNA microarray imputation.

PubMed

Jörnsten, Rebecka; Ouyang, Ming; Wang, Hui-Yu

2007-03-29

DNA microarray experiments are conducted in logical sets, such as time course profiling after a treatment is applied to the samples, or comparisons of the samples under two or more conditions. Due to cost and design constraints of spotted cDNA microarray experiments, each logical set commonly includes only a small number of replicates per condition. Despite the vast improvement of the microarray technology in recent years, missing values are prevalent. Intuitively, imputation of missing values is best done using many replicates within the same logical set. In practice, there are few replicates and thus reliable imputation within logical sets is difficult. However, it is in the case of few replicates that the presence of missing values, and how they are imputed, can have the most profound impact on the outcome of downstream analyses (e.g. significance analysis and clustering). This study explores the feasibility of imputation across logical sets, using the vast amount of publicly available microarray data to improve imputation reliability in the small sample size setting. We download all cDNA microarray data of Saccharomyces cerevisiae, Arabidopsis thaliana, and Caenorhabditis elegans from the Stanford Microarray Database. Through cross-validation and simulation, we find that, for all three species, our proposed imputation using data from public databases is far superior to imputation within a logical set, sometimes to an astonishing degree. Furthermore, the imputation root mean square error for significant genes is generally a lot less than that of non-significant ones. Since downstream analysis of significant genes, such as clustering and network analysis, can be very sensitive to small perturbations of estimated gene effects, it is highly recommended that researchers apply reliable data imputation prior to further analysis. Our method can also be applied to cDNA microarray experiments from other species, provided good reference data are available.

Inter-laboratory comparison of xenobiotic clearance rates determined using cryopreserved trout hepatocytes for improving bioaccumulation predictions

EPA Science Inventory

Hepatic biotransformation is an important determinant of chemical bioaccumulation in fish. Consequently, bioaccumulation models can be improved using estimates of chemical biotransformation rates. Cryopreserved trout hepatocytes have been used to measure the clearance rates of so...

Inter-laboratory comparison of clearance rates of xenobiotics by cryopreserved trout hepatocytes for the prediction of bioaccumulation potential

EPA Science Inventory

Hepatic biotransformation is an important determinant of chemical bioaccumulation in fish. Consequently, improvements to bioaccumulation models can be made using estimates of chemical biotransformation rates. Cryopreserved trout hepatocytes have previously been used to measure ...

Development and initial evaluation of a reconstituted water formulation that better represents natural waters(poster)

EPA Science Inventory

The use of reconstituted waters is deeply entrenched in many standardized aquatic toxicity testing protocols The primary appeal of reconstituted waters is inter-laboratory comparability, such that experiments performed in different laboratories can be conducted in (nominally) id...

Development and Initial Evaluation of a Reconstituted Water Formulation that Better Represents Natural Waters

EPA Science Inventory

The use of reconstituted waters is deeply entrenched in many standardized aquatic toxicity testing protocols. The primary appeal of reconstituted waters is inter-laboratory comparability, such that experiments performed in different laboratories can be conducted in (nominally) id...

INTERLABORATORY METHODS COMPARISON FOR THE TOTAL ORGANIC CARBON ANALYSIS OF AQUIFER MATERIALS

EPA Science Inventory

The total organic carbon (TOC) content of aquifer materials has been found to have significant effects on the movement of pollutants in the subsurface environment. Accurate quantification of TOC is therefore of great im- portance to research in groundwater contamination. However,...

9 CFR 439.51 - Probation of accreditation.

Code of Federal Regulations, 2010 CFR

2010-01-01

... ACCREDITATION OF NON-FEDERAL CHEMISTRY LABORATORIES § 439.51 Probation of accreditation. Upon a determination by the Administrator, a laboratory will be placed on probation for the following reasons: (a) If the laboratory fails to complete more than one interlaboratory accreditation maintenance check sample analysis as...

Certification by the Karl Fischer method of the water content in SRM 2890, Water Saturated 1-Octanol, and the analysis of associated interlaboratory bias in the measurement process.

PubMed

Margolis, S A; Levenson, M

2000-05-01

The calibration of Karl Fischer instruments and reagents and the compensation for instrumental bias are essential to the accurate measurement of trace levels of water in organic and inorganic chemicals. A stable, nonhygroscopic standard, Water Saturated Octanol, which is compatible with the Karl Fischer reagents, has been prepared. This material, Standard Reference Material (SRM) 2890, is homogeneous and is certified to contain 39.24 +/- 0.85 mg water/mL (expanded uncertainty) of solution (47.3 +/- 1.0 mg water/g solution, expanded uncertainty) at 21.5 degrees C. The solubility of water in -octanol has been shown to be nearly constant between 10 degrees C and 30 degrees C (i.e., within 1% of the value at 21.5 degrees C). The results of an interlaboratory comparison exercise illustrate the utility of SRM 2890 in assessing the accuracy and bias of Karl Fischer instruments and measurements.

Determination of Minor and Trace Metals in Aluminum and Aluminum Alloys by ICP-AES; Evaluation of the Uncertainty and Limit of Quantitation from Interlaboratory Testing.

PubMed

Uemoto, Michihisa; Makino, Masanori; Ota, Yuji; Sakaguchi, Hiromi; Shimizu, Yukari; Sato, Kazuhiro

2018-01-01

Minor and trace metals in aluminum and aluminum alloys have been determined by inductively coupled plasma atomic emission spectrometry (ICP-AES) as an interlaboratory testing toward standardization. The trueness of the measured data was successfully investigated to improve the analytical protocols, using certified reference materials of aluminum. Their precision could also be evaluated, feasible to estimate the uncertainties separately. The accuracy (trueness and precision) of the data were finally in good agreement with the certified values and assigned uncertainties. Repeated measurements of aluminum solutions with different concentrations of the analytes revealed the relative standard deviations of the measurements with concentrations, thus enabling their limits of quantitation. They differed separately and also showed slightly higher values with an aluminum matrix than those without one. In addition, the upper limit of the detectable concentration of silicon with simple acid digestion was estimated to be 0.03 % in the mass fraction.

Forensic interlaboratory evaluation of the ForFLUID kit for vaginal fluids identification.

PubMed

Giampaoli, Saverio; Alessandrini, Federica; Berti, Andrea; Ripani, Luigi; Choi, Ajin; Crab, Roselien; De Vittori, Elisabetta; Egyed, Balazs; Haas, Cordula; Lee, Hwan Young; Korabecná, Marie; Noel, Fabrice; Podini, Daniele; Tagliabracci, Adriano; Valentini, Alessio; Romano Spica, Vincenzo

2014-01-01

Identification of vaginal fluids is an important step in the process of sexual assaults confirmation. Advances in both microbiology and molecular biology defined technical approaches allowing the discrimination of body fluids. These protocols are based on the identification of specific bacterial communities by microfloraDNA (mfDNA) amplification. A multiplex real time-PCR assay (ForFLUID kit) has been developed for identifying biological fluids and for discrimination among vaginal, oral and fecal samples. In order to test its efficacy and reliability of the assay in the identification of vaginal fluids, an interlaboratory evaluation has been performed on homogeneous vaginal swabs. All the involved laboratories were able to correctly recognize all the vaginal swabs, and no false positives were identified when the assay was applied on non-vaginal samples. The assay represents an useful molecular tool that can be easily adopted by forensic geneticists involved in vaginal fluid identification. Copyright © 2013 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.

Finding Patterns of Emergence in Science and Technology

DTIC Science & Technology

2012-09-24

formal evaluation scheduled – Case Studies, Eight Examples: Tissue Engineering, Cold Fusion, RF Metamaterials, DNA Microarrays, Genetic Algorithms, RNAi...emerging capabilities Case Studies, Eight Examples: • Tissue Engineering, Cold Fusion, RF Metamaterials, DNA Microarrays, Genetic Algorithms...Evidence Quality (i.e., the rubric ) and deliver comprehensible evidential support for nomination • Demonstrate proof-of-concept nomination for Chinese

Systematic Omics Analysis Review (SOAR) Tool to Support Risk Assessment

PubMed Central

McConnell, Emma R.; Bell, Shannon M.; Cote, Ila; Wang, Rong-Lin; Perkins, Edward J.; Garcia-Reyero, Natàlia; Gong, Ping; Burgoon, Lyle D.

2014-01-01

Environmental health risk assessors are challenged to understand and incorporate new data streams as the field of toxicology continues to adopt new molecular and systems biology technologies. Systematic screening reviews can help risk assessors and assessment teams determine which studies to consider for inclusion in a human health assessment. A tool for systematic reviews should be standardized and transparent in order to consistently determine which studies meet minimum quality criteria prior to performing in-depth analyses of the data. The Systematic Omics Analysis Review (SOAR) tool is focused on assisting risk assessment support teams in performing systematic reviews of transcriptomic studies. SOAR is a spreadsheet tool of 35 objective questions developed by domain experts, focused on transcriptomic microarray studies, and including four main topics: test system, test substance, experimental design, and microarray data. The tool will be used as a guide to identify studies that meet basic published quality criteria, such as those defined by the Minimum Information About a Microarray Experiment standard and the Toxicological Data Reliability Assessment Tool. Seven scientists were recruited to test the tool by using it to independently rate 15 published manuscripts that study chemical exposures with microarrays. Using their feedback, questions were weighted based on importance of the information and a suitability cutoff was set for each of the four topic sections. The final validation resulted in 100% agreement between the users on four separate manuscripts, showing that the SOAR tool may be used to facilitate the standardized and transparent screening of microarray literature for environmental human health risk assessment. PMID:25531884

Optimized Probe Masking for Comparative Transcriptomics of Closely Related Species

PubMed Central

Poeschl, Yvonne; Delker, Carolin; Trenner, Jana; Ullrich, Kristian Karsten; Quint, Marcel; Grosse, Ivo

2013-01-01

Microarrays are commonly applied to study the transcriptome of specific species. However, many available microarrays are restricted to model organisms, and the design of custom microarrays for other species is often not feasible. Hence, transcriptomics approaches of non-model organisms as well as comparative transcriptomics studies among two or more species often make use of cost-intensive RNAseq studies or, alternatively, by hybridizing transcripts of a query species to a microarray of a closely related species. When analyzing these cross-species microarray expression data, differences in the transcriptome of the query species can cause problems, such as the following: (i) lower hybridization accuracy of probes due to mismatches or deletions, (ii) probes binding multiple transcripts of different genes, and (iii) probes binding transcripts of non-orthologous genes. So far, methods for (i) exist, but these neglect (ii) and (iii). Here, we propose an approach for comparative transcriptomics addressing problems (i) to (iii), which retains only transcript-specific probes binding transcripts of orthologous genes. We apply this approach to an Arabidopsis lyrata expression data set measured on a microarray designed for Arabidopsis thaliana, and compare it to two alternative approaches, a sequence-based approach and a genomic DNA hybridization-based approach. We investigate the number of retained probe sets, and we validate the resulting expression responses by qRT-PCR. We find that the proposed approach combines the benefit of sequence-based stringency and accuracy while allowing the expression analysis of much more genes than the alternative sequence-based approach. As an added benefit, the proposed approach requires probes to detect transcripts of orthologous genes only, which provides a superior base for biological interpretation of the measured expression responses. PMID:24260119

Improvement of experimental testing and network training conditions with genome-wide microarrays for more accurate predictions of drug gene targets

PubMed Central

2014-01-01

Background Genome-wide microarrays have been useful for predicting chemical-genetic interactions at the gene level. However, interpreting genome-wide microarray results can be overwhelming due to the vast output of gene expression data combined with off-target transcriptional responses many times induced by a drug treatment. This study demonstrates how experimental and computational methods can interact with each other, to arrive at more accurate predictions of drug-induced perturbations. We present a two-stage strategy that links microarray experimental testing and network training conditions to predict gene perturbations for a drug with a known mechanism of action in a well-studied organism. Results S. cerevisiae cells were treated with the antifungal, fluconazole, and expression profiling was conducted under different biological conditions using Affymetrix genome-wide microarrays. Transcripts were filtered with a formal network-based method, sparse simultaneous equation models and Lasso regression (SSEM-Lasso), under different network training conditions. Gene expression results were evaluated using both gene set and single gene target analyses, and the drug’s transcriptional effects were narrowed first by pathway and then by individual genes. Variables included: (i) Testing conditions – exposure time and concentration and (ii) Network training conditions – training compendium modifications. Two analyses of SSEM-Lasso output – gene set and single gene – were conducted to gain a better understanding of how SSEM-Lasso predicts perturbation targets. Conclusions This study demonstrates that genome-wide microarrays can be optimized using a two-stage strategy for a more in-depth understanding of how a cell manifests biological reactions to a drug treatment at the transcription level. Additionally, a more detailed understanding of how the statistical model, SSEM-Lasso, propagates perturbations through a network of gene regulatory interactions is achieved. PMID:24444313

Development and experimental validation of a 20K Atlantic cod (Gadus morhua) oligonucleotide microarray based on a collection of over 150,000 ESTs.

PubMed

Booman, Marije; Borza, Tudor; Feng, Charles Y; Hori, Tiago S; Higgins, Brent; Culf, Adrian; Léger, Daniel; Chute, Ian C; Belkaid, Anissa; Rise, Marlies; Gamperl, A Kurt; Hubert, Sophie; Kimball, Jennifer; Ouellette, Rodney J; Johnson, Stewart C; Bowman, Sharen; Rise, Matthew L

2011-08-01

The collapse of Atlantic cod (Gadus morhua) wild populations strongly impacted the Atlantic cod fishery and led to the development of cod aquaculture. In order to improve aquaculture and broodstock quality, we need to gain knowledge of genes and pathways involved in Atlantic cod responses to pathogens and other stressors. The Atlantic Cod Genomics and Broodstock Development Project has generated over 150,000 expressed sequence tags from 42 cDNA libraries representing various tissues, developmental stages, and stimuli. We used this resource to develop an Atlantic cod oligonucleotide microarray containing 20,000 unique probes. Selection of sequences from the full range of cDNA libraries enables application of the microarray for a broad spectrum of Atlantic cod functional genomics studies. We included sequences that were highly abundant in suppression subtractive hybridization (SSH) libraries, which were enriched for transcripts responsive to pathogens or other stressors. These sequences represent genes that potentially play an important role in stress and/or immune responses, making the microarray particularly useful for studies of Atlantic cod gene expression responses to immune stimuli and other stressors. To demonstrate its value, we used the microarray to analyze the Atlantic cod spleen response to stimulation with formalin-killed, atypical Aeromonas salmonicida, resulting in a gene expression profile that indicates a strong innate immune response. These results were further validated by quantitative PCR analysis and comparison to results from previous analysis of an SSH library. This study shows that the Atlantic cod 20K oligonucleotide microarray is a valuable new tool for Atlantic cod functional genomics research.

Methodological study of affine transformations of gene expression data with proposed robust non-parametric multi-dimensional normalization method.

PubMed

Bengtsson, Henrik; Hössjer, Ola

2006-03-01

Low-level processing and normalization of microarray data are most important steps in microarray analysis, which have profound impact on downstream analysis. Multiple methods have been suggested to date, but it is not clear which is the best. It is therefore important to further study the different normalization methods in detail and the nature of microarray data in general. A methodological study of affine models for gene expression data is carried out. Focus is on two-channel comparative studies, but the findings generalize also to single- and multi-channel data. The discussion applies to spotted as well as in-situ synthesized microarray data. Existing normalization methods such as curve-fit ("lowess") normalization, parallel and perpendicular translation normalization, and quantile normalization, but also dye-swap normalization are revisited in the light of the affine model and their strengths and weaknesses are investigated in this context. As a direct result from this study, we propose a robust non-parametric multi-dimensional affine normalization method, which can be applied to any number of microarrays with any number of channels either individually or all at once. A high-quality cDNA microarray data set with spike-in controls is used to demonstrate the power of the affine model and the proposed normalization method. We find that an affine model can explain non-linear intensity-dependent systematic effects in observed log-ratios. Affine normalization removes such artifacts for non-differentially expressed genes and assures that symmetry between negative and positive log-ratios is obtained, which is fundamental when identifying differentially expressed genes. In addition, affine normalization makes the empirical distributions in different channels more equal, which is the purpose of quantile normalization, and may also explain why dye-swap normalization works or fails. All methods are made available in the aroma package, which is a platform-independent package for R.

A statistical model for investigating binding probabilities of DNA nucleotide sequences using microarrays.

PubMed

Lee, Mei-Ling Ting; Bulyk, Martha L; Whitmore, G A; Church, George M

2002-12-01

There is considerable scientific interest in knowing the probability that a site-specific transcription factor will bind to a given DNA sequence. Microarray methods provide an effective means for assessing the binding affinities of a large number of DNA sequences as demonstrated by Bulyk et al. (2001, Proceedings of the National Academy of Sciences, USA 98, 7158-7163) in their study of the DNA-binding specificities of Zif268 zinc fingers using microarray technology. In a follow-up investigation, Bulyk, Johnson, and Church (2002, Nucleic Acid Research 30, 1255-1261) studied the interdependence of nucleotides on the binding affinities of transcription proteins. Our article is motivated by this pair of studies. We present a general statistical methodology for analyzing microarray intensity measurements reflecting DNA-protein interactions. The log probability of a protein binding to a DNA sequence on an array is modeled using a linear ANOVA model. This model is convenient because it employs familiar statistical concepts and procedures and also because it is effective for investigating the probability structure of the binding mechanism.

Analysis of developmental gene conservation in the Actinomycetales using DNA/DNA microarray comparisons.

PubMed

Kirby, Ralph; Herron, Paul; Hoskisson, Paul

2011-02-01

Based on available genome sequences, Actinomycetales show significant gene synteny across a wide range of species and genera. In addition, many genera show varying degrees of complex morphological development. Using the presence of gene synteny as a basis, it is clear that an analysis of gene conservation across the Streptomyces and various other Actinomycetales will provide information on both the importance of genes and gene clusters and the evolution of morphogenesis in these bacteria. Genome sequencing, although becoming cheaper, is still relatively expensive for comparing large numbers of strains. Thus, a heterologous DNA/DNA microarray hybridization dataset based on a Streptomyces coelicolor microarray allows a cheaper and greater depth of analysis of gene conservation. This study, using both bioinformatical and microarray approaches, was able to classify genes previously identified as involved in morphogenesis in Streptomyces into various subgroups in terms of conservation across species and genera. This will allow the targeting of genes for further study based on their importance at the species level and at higher evolutionary levels.

Cross species analysis of microarray expression data

PubMed Central

Lu, Yong; Huggins, Peter; Bar-Joseph, Ziv

2009-01-01

Motivation: Many biological systems operate in a similar manner across a large number of species or conditions. Cross-species analysis of sequence and interaction data is often applied to determine the function of new genes. In contrast to these static measurements, microarrays measure the dynamic, condition-specific response of complex biological systems. The recent exponential growth in microarray expression datasets allows researchers to combine expression experiments from multiple species to identify genes that are not only conserved in sequence but also operated in a similar way in the different species studied. Results: In this review we discuss the computational and technical challenges associated with these studies, the approaches that have been developed to address these challenges and the advantages of cross-species analysis of microarray data. We show how successful application of these methods lead to insights that cannot be obtained when analyzing data from a single species. We also highlight current open problems and discuss possible ways to address them. Contact: zivbj@cs.cmu.edu PMID:19357096

Missing value imputation for microarray data: a comprehensive comparison study and a web tool.

PubMed

Chiu, Chia-Chun; Chan, Shih-Yao; Wang, Chung-Ching; Wu, Wei-Sheng

2013-01-01

Microarray data are usually peppered with missing values due to various reasons. However, most of the downstream analyses for microarray data require complete datasets. Therefore, accurate algorithms for missing value estimation are needed for improving the performance of microarray data analyses. Although many algorithms have been developed, there are many debates on the selection of the optimal algorithm. The studies about the performance comparison of different algorithms are still incomprehensive, especially in the number of benchmark datasets used, the number of algorithms compared, the rounds of simulation conducted, and the performance measures used. In this paper, we performed a comprehensive comparison by using (I) thirteen datasets, (II) nine algorithms, (III) 110 independent runs of simulation, and (IV) three types of measures to evaluate the performance of each imputation algorithm fairly. First, the effects of different types of microarray datasets on the performance of each imputation algorithm were evaluated. Second, we discussed whether the datasets from different species have different impact on the performance of different algorithms. To assess the performance of each algorithm fairly, all evaluations were performed using three types of measures. Our results indicate that the performance of an imputation algorithm mainly depends on the type of a dataset but not on the species where the samples come from. In addition to the statistical measure, two other measures with biological meanings are useful to reflect the impact of missing value imputation on the downstream data analyses. Our study suggests that local-least-squares-based methods are good choices to handle missing values for most of the microarray datasets. In this work, we carried out a comprehensive comparison of the algorithms for microarray missing value imputation. Based on such a comprehensive comparison, researchers could choose the optimal algorithm for their datasets easily. Moreover, new imputation algorithms could be compared with the existing algorithms using this comparison strategy as a standard protocol. In addition, to assist researchers in dealing with missing values easily, we built a web-based and easy-to-use imputation tool, MissVIA (http://cosbi.ee.ncku.edu.tw/MissVIA), which supports many imputation algorithms. Once users upload a real microarray dataset and choose the imputation algorithms, MissVIA will determine the optimal algorithm for the users' data through a series of simulations, and then the imputed results can be downloaded for the downstream data analyses.

Hybrid genetic algorithm-neural network: feature extraction for unpreprocessed microarray data.

PubMed

Tong, Dong Ling; Schierz, Amanda C

2011-09-01

Suitable techniques for microarray analysis have been widely researched, particularly for the study of marker genes expressed to a specific type of cancer. Most of the machine learning methods that have been applied to significant gene selection focus on the classification ability rather than the selection ability of the method. These methods also require the microarray data to be preprocessed before analysis takes place. The objective of this study is to develop a hybrid genetic algorithm-neural network (GANN) model that emphasises feature selection and can operate on unpreprocessed microarray data. The GANN is a hybrid model where the fitness value of the genetic algorithm (GA) is based upon the number of samples correctly labelled by a standard feedforward artificial neural network (ANN). The model is evaluated by using two benchmark microarray datasets with different array platforms and differing number of classes (a 2-class oligonucleotide microarray data for acute leukaemia and a 4-class complementary DNA (cDNA) microarray dataset for SRBCTs (small round blue cell tumours)). The underlying concept of the GANN algorithm is to select highly informative genes by co-evolving both the GA fitness function and the ANN weights at the same time. The novel GANN selected approximately 50% of the same genes as the original studies. This may indicate that these common genes are more biologically significant than other genes in the datasets. The remaining 50% of the significant genes identified were used to build predictive models and for both datasets, the models based on the set of genes extracted by the GANN method produced more accurate results. The results also suggest that the GANN method not only can detect genes that are exclusively associated with a single cancer type but can also explore the genes that are differentially expressed in multiple cancer types. The results show that the GANN model has successfully extracted statistically significant genes from the unpreprocessed microarray data as well as extracting known biologically significant genes. We also show that assessing the biological significance of genes based on classification accuracy may be misleading and though the GANN's set of extra genes prove to be more statistically significant than those selected by other methods, a biological assessment of these genes is highly recommended to confirm their functionality. Copyright © 2011 Elsevier B.V. All rights reserved.

Hybridization chain reaction amplification for highly sensitive fluorescence detection of DNA with dextran coated microarrays.

PubMed

Chao, Jie; Li, Zhenhua; Li, Jing; Peng, Hongzhen; Su, Shao; Li, Qian; Zhu, Changfeng; Zuo, Xiaolei; Song, Shiping; Wang, Lianhui; Wang, Lihua

2016-07-15

Microarrays of biomolecules hold great promise in the fields of genomics, proteomics, and clinical assays on account of their remarkably parallel and high-throughput assay capability. However, the fluorescence detection used in most conventional DNA microarrays is still limited by sensitivity. In this study, we have demonstrated a novel universal and highly sensitive platform for fluorescent detection of sequence specific DNA at the femtomolar level by combining dextran-coated microarrays with hybridization chain reaction (HCR) signal amplification. Three-dimensional dextran matrix was covalently coated on glass surface as the scaffold to immobilize DNA recognition probes to increase the surface binding capacity and accessibility. DNA nanowire tentacles were formed on the matrix surface for efficient signal amplification by capturing multiple fluorescent molecules in a highly ordered way. By quantifying microscopic fluorescent signals, the synergetic effects of dextran and HCR greatly improved sensitivity of DNA microarrays, with a detection limit of 10fM (1×10(5) molecules). This detection assay could recognize one-base mismatch with fluorescence signals dropped down to ~20%. This cost-effective microarray platform also worked well with samples in serum and thus shows great potential for clinical diagnosis. Copyright © 2016 Elsevier B.V. All rights reserved.

Controlling false-negative errors in microarray differential expression analysis: a PRIM approach.

PubMed

Cole, Steve W; Galic, Zoran; Zack, Jerome A

2003-09-22

Theoretical considerations suggest that current microarray screening algorithms may fail to detect many true differences in gene expression (Type II analytic errors). We assessed 'false negative' error rates in differential expression analyses by conventional linear statistical models (e.g. t-test), microarray-adapted variants (e.g. SAM, Cyber-T), and a novel strategy based on hold-out cross-validation. The latter approach employs the machine-learning algorithm Patient Rule Induction Method (PRIM) to infer minimum thresholds for reliable change in gene expression from Boolean conjunctions of fold-induction and raw fluorescence measurements. Monte Carlo analyses based on four empirical data sets show that conventional statistical models and their microarray-adapted variants overlook more than 50% of genes showing significant up-regulation. Conjoint PRIM prediction rules recover approximately twice as many differentially expressed transcripts while maintaining strong control over false-positive (Type I) errors. As a result, experimental replication rates increase and total analytic error rates decline. RT-PCR studies confirm that gene inductions detected by PRIM but overlooked by other methods represent true changes in mRNA levels. PRIM-based conjoint inference rules thus represent an improved strategy for high-sensitivity screening of DNA microarrays. Freestanding JAVA application at http://microarray.crump.ucla.edu/focus

Strategies for comparing gene expression profiles from different microarray platforms: application to a case-control experiment.

PubMed

Severgnini, Marco; Bicciato, Silvio; Mangano, Eleonora; Scarlatti, Francesca; Mezzelani, Alessandra; Mattioli, Michela; Ghidoni, Riccardo; Peano, Clelia; Bonnal, Raoul; Viti, Federica; Milanesi, Luciano; De Bellis, Gianluca; Battaglia, Cristina

2006-06-01

Meta-analysis of microarray data is increasingly important, considering both the availability of multiple platforms using disparate technologies and the accumulation in public repositories of data sets from different laboratories. We addressed the issue of comparing gene expression profiles from two microarray platforms by devising a standardized investigative strategy. We tested this procedure by studying MDA-MB-231 cells, which undergo apoptosis on treatment with resveratrol. Gene expression profiles were obtained using high-density, short-oligonucleotide, single-color microarray platforms: GeneChip (Affymetrix) and CodeLink (Amersham). Interplatform analyses were carried out on 8414 common transcripts represented on both platforms, as identified by LocusLink ID, representing 70.8% and 88.6% of annotated GeneChip and CodeLink features, respectively. We identified 105 differentially expressed genes (DEGs) on CodeLink and 42 DEGs on GeneChip. Among them, only 9 DEGs were commonly identified by both platforms. Multiple analyses (BLAST alignment of probes with target sequences, gene ontology, literature mining, and quantitative real-time PCR) permitted us to investigate the factors contributing to the generation of platform-dependent results in single-color microarray experiments. An effective approach to cross-platform comparison involves microarrays of similar technologies, samples prepared by identical methods, and a standardized battery of bioinformatic and statistical analyses.

Living Cell Microarrays: An Overview of Concepts

PubMed Central

Jonczyk, Rebecca; Kurth, Tracy; Lavrentieva, Antonina; Walter, Johanna-Gabriela; Scheper, Thomas; Stahl, Frank

2016-01-01

Living cell microarrays are a highly efficient cellular screening system. Due to the low number of cells required per spot, cell microarrays enable the use of primary and stem cells and provide resolution close to the single-cell level. Apart from a variety of conventional static designs, microfluidic microarray systems have also been established. An alternative format is a microarray consisting of three-dimensional cell constructs ranging from cell spheroids to cells encapsulated in hydrogel. These systems provide an in vivo-like microenvironment and are preferably used for the investigation of cellular physiology, cytotoxicity, and drug screening. Thus, many different high-tech microarray platforms are currently available. Disadvantages of many systems include their high cost, the requirement of specialized equipment for their manufacture, and the poor comparability of results between different platforms. In this article, we provide an overview of static, microfluidic, and 3D cell microarrays. In addition, we describe a simple method for the printing of living cell microarrays on modified microscope glass slides using standard DNA microarray equipment available in most laboratories. Applications in research and diagnostics are discussed, e.g., the selective and sensitive detection of biomarkers. Finally, we highlight current limitations and the future prospects of living cell microarrays. PMID:27600077

An interlaboratory comparison study on the measurement of elements in PM10

NASA Astrophysics Data System (ADS)

Yatkin, Sinan; Belis, Claudio A.; Gerboles, Michel; Calzolai, Giulia; Lucarelli, Franco; Cavalli, Fabrizia; Trzepla, Krystyna

2016-01-01

An inter-laboratory comparison study was conducted to measure elemental loadings on PM10 samples, collected in Ispra, a regional background/rural site in Italy, using three different XRF (X-ray Fluorescence) methods, namely Epsilon 5 by linear calibration, Quant'X by the standardless analysis, and PIXE (Particle Induced X-ray Emission) with linear calibration. A subset of samples was also analyzed by ICP-MS (Inductively Coupled Plasma-Mass Spectrometry). Several metrics including method detection limits (MDLs), precision, bias from a NIST standard reference material (SRM 2783) quoted values, relative absolute difference, orthogonal regression and the ratio of the absolute difference between the methods to claimed uncertainty were used to compare the laboratories. The MDLs were found to be comparable for many elements. Precision estimates were less than 10% for the majority of the elements. Absolute biases from SRM 2783 remained less than 20% for the majority of certified elements. The regression results of PM10 samples showed that the three XRF laboratories measured very similar mass loadings for S, K, Ti, Mn, Fe, Cu, Br, Sr and Pb with slopes within 20% of unity. The ICP-MS results confirmed the agreement and discrepancies between XRF laboratories for Al, K, Ca, Ti, V, Cu, Sr and Pb. The ICP-MS results are inconsistent with the XRF laboratories for Fe and Zn. The absolute differences between the XRF laboratories generally remained within their claimed uncertainties, showing a pattern generally consistent with the orthogonal regression results.

Harmonisation of seven common enzyme results through EQA.

PubMed

Weykamp, Cas; Franck, Paul; Gunnewiek, Jacqueline Klein; de Jonge, Robert; Kuypers, Aldy; van Loon, Douwe; Steigstra, Herman; Cobbaert, Christa

2014-11-01

Equivalent results between different laboratories enable optimal patient care and can be achieved with harmonisation. We report on EQA-initiated national harmonisation of seven enzymes using commutable samples. EQA samples were prepared from human serum spiked with human recombinant enzymes. Target values were assigned with the IFCC Reference Measurement Procedures. The same samples were included at four occasions in the EQA programmes of 2012 and 2013. Laboratories were encouraged to report IFCC traceable results. A parallel study was done to confirm commutability of the samples. Of the 223 participating laboratories, 95% reported IFCC traceable results, ranging from 98% (ASAT) to 87% (amylase). Users of Roche and Siemens (97%) more frequently reported in IFCC traceable results than users of Abbott (91%), Beckman (90%), and Olympus (87%). The success of harmonisation, expressed as the recovery of assigned values and the inter-laboratory CV was: ALAT (recovery 100%; inter-lab CV 4%), ASAT (102%; 4%), LD (98%; 3%), CK (101%; 5%), GGT (98%; 4%), AP (96%; 6%), amylase (99%; 4%). There were no significant differences between the manufacturers. Commutability was demonstrated in the parallel study. Equal results in the same sample in the 2012 and 2013 EQA programmes demonstrated stability of the samples. The EQA-initiated national harmonisation of seven enzymes, using stable, commutable human serum samples, spiked with human recombinant enzymes, and targeted with the IFCC Reference Measurement Procedures, was successful in terms of implementation of IFCC traceable results (95%), recovery of the target (99%), and inter-laboratory CV (4%).

Harmonization of urine albumin/creatinine ratio (ACR) results: a study based on an external quality assessment program in Polish laboratories.

PubMed

Ćwiklińska, Agnieszka; Dąbrowska, Hanna; Kowalski, Robert; Kuchta, Agnieszka; Kortas-Stempak, Barbara; Fijałkowska, Aleksandra; Bednarczuk, Gabriela; Jankowski, Maciej

2018-05-11

The ratio of albumin to creatinine (ACR) is an important parameter used for detection of albuminuria in patients with early kidney damage. The aim of the study was to evaluate the harmonization of ACR results among Polish participants in an international external quality assessment (EQA) program, and to evaluate the impact of albumin and creatinine analytical performance on the harmonization of ACR results. We analyzed 182 results of albumin, 202 of creatinine, and 180 of ACR obtained from Polish laboratories in an EQA program organized by Labquality. The dispersion of the results in surveys and percentage differences between the results and target values were calculated. Moreover, differences between method groups were assessed. The inter-laboratory coefficient of variation (CV) for ACR was 36%. Only 74% of results of Polish laboratories were within the target limits; for 11% of the results, an incorrect albuminuria category would have been reported. The inter-laboratory CV for albumin was 20%, 2.6-fold higher than for creatinine. Significant differences between method groups for albumin determination have been observed, even when the same measurement technique was used. The greatest difference between two groups was 23%, 2.5-fold greater in comparison to creatinine. There is an insufficient harmonization of ACR values among Polish laboratories, caused mainly by urine albumin analytical performance. Given the important role of ACR in the classification, monitoring and treatment of kidney damage, the harmonization of albumin measurements is crucial and urgently needed.

Interlaboratory variability in screening for Bonamia ostreae, a protistan parasite of the European flat oyster Ostrea edulis.

PubMed

Flannery, Grace; Lynch, Sharon A; Longshaw, Matt; Stone, David; Martin, Paul; Ramilo, Andrea; Villalba, Antonio; Culloty, Sarah C

2014-07-24

The spread of the protozoan parasite Bonamia ostreae is of major concern to the European flat oyster Ostrea edulis industry. Many studies have looked at the sensitivity of individual methods available to screen for B. ostreae, but in this study, 3 separate laboratories examined 4 methods of diagnosis currently used routinely in laboratories: heart imprints, histology, polymerase chain reaction (PCR) and in situ hybridisation (ISH). The results were compared to estimate interlaboratory variability. Heart imprints and histology had the highest reproducibility amongst the 3 laboratories, with greatest agreement between detection of infected and uninfected individuals. PCR had the highest detection level in every laboratory. These positives were related to the presence of confirmed infections but also in unconfirmed infections, possibly due to the presence of traces of B. ostreae DNA in oysters where clinical infections were not observed. PCR, in combination with histology or ISH, provided the most reliable detection levels in every laboratory. Variation in results for PCR and ISH observed between laboratories may be due to the different protocols used by each laboratory for both methods. Overall, the findings from the 3 laboratories indicated that at least 2 methods, with fixed protocols, should be used for the accurate detection and determination of infection prevalence within a sample. This combination of methods would allow for a clearer and more precise diagnosis of B. ostreae, preventing further spread of the disease and providing more accurate detection levels and epidemiological information.

Estimation of discrimination errors in the technique for determining the geographic origin of onions by mineral composition: interlaboratory study.

PubMed

Ariyama, Kaoru; Kadokura, Masashi; Suzuki, Tadanao

2008-01-01

Techniques to determine the geographic origin of foods have been developed for various agricultural and fishery products, and they have used various principles. Some of these techniques are already in use for checking the authenticity of the labeling. Many are based on multielement analysis and chemometrics. We have developed such a technique to determine the geographic origin of onions (Allium cepa L.). This technique, which determines whether an onion is from outside Japan, is designed for onions labeled as having a geographic origin of Hokkaido, Hyogo, or Saga, the main onion production areas in Japan. However, estimations of discrimination errors for this technique have not been fully conducted; they have been limited to those for discrimination models and do not include analytical errors. Interlaboratory studies were conducted to estimate the analytical errors of the technique. Four collaborators each determined 11 elements (Na, Mg, P, Mn, Zn, Rb, Sr, Mo, Cd, Cs, and Ba) in 4 test materials of fresh and dried onions. Discrimination errors in this technique were estimated by summing (1) individual differences within lots, (2) variations between lots from the same production area, and (3) analytical errors. The discrimination errors for onions from Hokkaido, Hyogo, and Saga were estimated to be 2.3, 9.5, and 8.0%, respectively. Those for onions from abroad in determinations targeting Hokkaido, Hyogo, and Saga were estimated to be 28.2, 21.6, and 21.9%, respectively.

Evaluation of Cardiac Toxicity Biomarkers in Rats from Different Laboratories.

PubMed

Kim, Kyuri; Chini, Naseem; Fairchild, David G; Engle, Steven K; Reagan, William J; Summers, Sandra D; Mirsalis, Jon C

2016-12-01

There is a great need for improved diagnostic and prognostic accuracy of potential cardiac toxicity in drug development. This study reports the evaluation of several commercially available biomarker kits by 3 institutions (SRI, Eli Lilly, and Pfizer) for the discrimination between myocardial degeneration/necrosis and cardiac hypertrophy as well as the assessment of the interlaboratory and interplatform variation in results. Serum concentrations of natriuretic peptides (N-terminal pro-atrial natriuretic peptide [NT-proANP] and N-terminal pro-brain natriuretic peptide [NT-proBNP]), cardiac and skeletal troponins (cTnI, cTnT, and sTnI), myosin light chain 3 (Myl3), and fatty acid binding protein 3 (FABP3) were assessed in rats treated with minoxidil (MNX) and isoproterenol (ISO). MNX caused increased heart-to-body weight ratios and prominent elevations in NT-proANP and NT-proBNP concentrations detected at 24-hr postdose without elevation in troponins, Myl3, or FABP3 and with no abnormal histopathological findings. ISO caused ventricular leukocyte infiltration, myocyte fibrosis, and necrosis with increased concentrations of the natriuretic peptides, cardiac troponins, and Myl3. These results reinforce the advantages of a multimarker strategy in elucidating the underlying cause of cardiac insult and detecting myocardial tissue damage at 24-hr posttreatment. The interlaboratory and interplatform comparison analyses also showed that the data obtained from different laboratories and platforms are highly correlated and reproducible, making these biomarkers widely applicable in preclinical studies.

Identifying molecular features for prostate cancer with Gleason 7 based on microarray gene expression profiles.

PubMed

Bălăcescu, Loredana; Bălăcescu, O; Crişan, N; Fetica, B; Petruţ, B; Bungărdean, Cătălina; Rus, Meda; Tudoran, Oana; Meurice, G; Irimie, Al; Dragoş, N; Berindan-Neagoe, Ioana

2011-01-01

Prostate cancer represents the first leading cause of cancer among western male population, with different clinical behavior ranging from indolent to metastatic disease. Although many molecules and deregulated pathways are known, the molecular mechanisms involved in the development of prostate cancer are not fully understood. The aim of this study was to explore the molecular variation underlying the prostate cancer, based on microarray analysis and bioinformatics approaches. Normal and prostate cancer tissues were collected by macrodissection from prostatectomy pieces. All prostate cancer specimens used in our study were Gleason score 7. Gene expression microarray (Agilent Technologies) was used for Whole Human Genome evaluation. The bioinformatics and functional analysis were based on Limma and Ingenuity software. The microarray analysis identified 1119 differentially expressed genes between prostate cancer and normal prostate, which were up- or down-regulated at least 2-fold. P-values were adjusted for multiple testing using Benjamini-Hochberg method with a false discovery rate of 0.01. These genes were analyzed with Ingenuity Pathway Analysis software and were established 23 genetic networks. Our microarray results provide new information regarding the molecular networks in prostate cancer stratified as Gleason 7. These data highlighted gene expression profiles for better understanding of prostate cancer progression.

Detection of Alicyclobacillus species in fruit juice using a random genomic DNA microarray chip.

PubMed

Jang, Jun Hyeong; Kim, Sun-Joong; Yoon, Bo Hyun; Ryu, Jee-Hoon; Gu, Man Bock; Chang, Hyo-Ihl

2011-06-01

This study describes a method using a DNA microarray chip to rapidly and simultaneously detect Alicyclobacillus species in orange juice based on the hybridization of genomic DNA with random probes. Three food spoilage bacteria were used in this study: Alicyclobacillus acidocaldarius, Alicyclobacillus acidoterrestris, and Alicyclobacillus cycloheptanicus. The three Alicyclobacillus species were adjusted to 2 × 10(3) CFU/ml and inoculated into pasteurized 100% pure orange juice. Cy5-dCTP labeling was used for reference signals, and Cy3-dCTP was labeled for target genomic DNA. The molar ratio of 1:1 of Cy3-dCTP and Cy5-dCTP was used. DNA microarray chips were fabricated using randomly fragmented DNA of Alicyclobacillus spp. and were hybridized with genomic DNA extracted from Bacillus spp. Genomic DNA extracted from Alicyclobacillus spp. showed a significantly higher hybridization rate compared with DNA of Bacillus spp., thereby distinguishing Alicyclobacillus spp. from Bacillus spp. The results showed that the microarray DNA chip containing randomly fragmented genomic DNA was specific and clearly identified specific food spoilage bacteria. This microarray system is a good tool for rapid and specific detection of thermophilic spoilage bacteria, mainly Alicyclobacillus spp., and is useful and applicable to the fruit juice industry.

Analysis of host response to bacterial infection using error model based gene expression microarray experiments

PubMed Central

Stekel, Dov J.; Sarti, Donatella; Trevino, Victor; Zhang, Lihong; Salmon, Mike; Buckley, Chris D.; Stevens, Mark; Pallen, Mark J.; Penn, Charles; Falciani, Francesco

2005-01-01

A key step in the analysis of microarray data is the selection of genes that are differentially expressed. Ideally, such experiments should be properly replicated in order to infer both technical and biological variability, and the data should be subjected to rigorous hypothesis tests to identify the differentially expressed genes. However, in microarray experiments involving the analysis of very large numbers of biological samples, replication is not always practical. Therefore, there is a need for a method to select differentially expressed genes in a rational way from insufficiently replicated data. In this paper, we describe a simple method that uses bootstrapping to generate an error model from a replicated pilot study that can be used to identify differentially expressed genes in subsequent large-scale studies on the same platform, but in which there may be no replicated arrays. The method builds a stratified error model that includes array-to-array variability, feature-to-feature variability and the dependence of error on signal intensity. We apply this model to the characterization of the host response in a model of bacterial infection of human intestinal epithelial cells. We demonstrate the effectiveness of error model based microarray experiments and propose this as a general strategy for a microarray-based screening of large collections of biological samples. PMID:15800204

An international standardization programme towards the application of gene expression profiling in routine leukaemia diagnostics: the Microarray Innovations in LEukemia study prephase

PubMed Central

Kohlmann, Alexander; Kipps, Thomas J; Rassenti, Laura Z; Downing, James R; Shurtleff, Sheila A; Mills, Ken I; Gilkes, Amanda F; Hofmann, Wolf-Karsten; Basso, Giuseppe; Dell’Orto, Marta Campo; Foà, Robin; Chiaretti, Sabina; De Vos, John; Rauhut, Sonja; Papenhausen, Peter R; Hernández, Jesus M; Lumbreras, Eva; Yeoh, Allen E; Koay, Evelyn S; Li, Rachel; Liu, Wei-min; Williams, Paul M; Wieczorek, Lothar; Haferlach, Torsten

2008-01-01

Gene expression profiling has the potential to enhance current methods for the diagnosis of haematological malignancies. Here, we present data on 204 analyses from an international standardization programme that was conducted in 11 laboratories as a prephase to the Microarray Innovations in LEukemia (MILE) study. Each laboratory prepared two cell line samples, together with three replicate leukaemia patient lysates in two distinct stages: (i) a 5-d course of protocol training, and (ii) independent proficiency testing. Unsupervised, supervised, and r2 correlation analyses demonstrated that microarray analysis can be performed with remarkably high intra-laboratory reproducibility and with comparable quality and reliability. PMID:18573112

ELISA-BASE: An Integrated Bioinformatics Tool for Analyzing and Tracking ELISA Microarray Data

DOE Office of Scientific and Technical Information (OSTI.GOV)

White, Amanda M.; Collett, James L.; Seurynck-Servoss, Shannon L.

ELISA-BASE is an open-source database for capturing, organizing and analyzing protein enzyme-linked immunosorbent assay (ELISA) microarray data. ELISA-BASE is an extension of the BioArray Soft-ware Environment (BASE) database system, which was developed for DNA microarrays. In order to make BASE suitable for protein microarray experiments, we developed several plugins for importing and analyzing quantitative ELISA microarray data. Most notably, our Protein Microarray Analysis Tool (ProMAT) for processing quantita-tive ELISA data is now available as a plugin to the database.

In vitro to in vivo extrapolation of hepatic metabolism in fish: An inter-laboratory comparison of in vitro methods - presentation

EPA Science Inventory

Chemical biotransformation represents the largest source of uncertainty in chemical bioaccumulation assessments. Model-based estimates of chemical bioconcentration in fish may be greatly improved by including biotransformation rates, as measured in vitro. Substrate depletion assa...

Development of micro-electrode array based tests for neurotoxicity: assessment of interlaboratory reproducibility with neuroactive chemicals

EPA Science Inventory

Neuronal assemblies within the Central Nervous System (CNS) produce spontaneous or stimulus-evoked electrophysiological activity that can be monitored and quantified in terms of action potential patterns. Such patterns provide a sensitive endpoint to detect effects of chemicals, ...

Thermodynamically optimal whole-genome tiling microarray design and validation.

PubMed

Cho, Hyejin; Chou, Hui-Hsien

2016-06-13

Microarray is an efficient apparatus to interrogate the whole transcriptome of species. Microarray can be designed according to annotated gene sets, but the resulted microarrays cannot be used to identify novel transcripts and this design method is not applicable to unannotated species. Alternatively, a whole-genome tiling microarray can be designed using only genomic sequences without gene annotations, and it can be used to detect novel RNA transcripts as well as known genes. The difficulty with tiling microarray design lies in the tradeoff between probe-specificity and coverage of the genome. Sequence comparison methods based on BLAST or similar software are commonly employed in microarray design, but they cannot precisely determine the subtle thermodynamic competition between probe targets and partially matched probe nontargets during hybridizations. Using the whole-genome thermodynamic analysis software PICKY to design tiling microarrays, we can achieve maximum whole-genome coverage allowable under the thermodynamic constraints of each target genome. The resulted tiling microarrays are thermodynamically optimal in the sense that all selected probes share the same melting temperature separation range between their targets and closest nontargets, and no additional probes can be added without violating the specificity of the microarray to the target genome. This new design method was used to create two whole-genome tiling microarrays for Escherichia coli MG1655 and Agrobacterium tumefaciens C58 and the experiment results validated the design.

Use of Network Inference to Elucidate Common and Chemical-specific Effects on Steoidogenesis

EPA Science Inventory

Microarray data is a key source for modeling gene regulatory interactions. Regulatory network models based on multiple datasets are potentially more robust and can provide greater confidence. In this study, we used network modeling on microarray data generated by exposing the fat...

Diagnostic Yield of Chromosomal Microarray Analysis in a Cohort of Patients with Autism Spectrum Disorders from a Highly Consanguineous Population

ERIC Educational Resources Information Center

Al-Mamari, Watfa; Al-Saegh, Abeer; Al-Kindy, Adila; Bruwer, Zandre; Al-Murshedi, Fathiya; Al-Thihli, Khalid

2015-01-01

Autism Spectrum Disorders are a complicated group of disorders characterized with heterogeneous genetic etiologies. The genetic investigations for this group of disorders have expanded considerably over the past decade. In our study we designed a tired approach and studied the diagnostic yield of chromosomal microarray analysis on patients…

Apparently low reproducibility of true differential expression discoveries in microarray studies.

PubMed

Zhang, Min; Yao, Chen; Guo, Zheng; Zou, Jinfeng; Zhang, Lin; Xiao, Hui; Wang, Dong; Yang, Da; Gong, Xue; Zhu, Jing; Li, Yanhui; Li, Xia

2008-09-15

Differentially expressed gene (DEG) lists detected from different microarray studies for a same disease are often highly inconsistent. Even in technical replicate tests using identical samples, DEG detection still shows very low reproducibility. It is often believed that current small microarray studies will largely introduce false discoveries. Based on a statistical model, we show that even in technical replicate tests using identical samples, it is highly likely that the selected DEG lists will be very inconsistent in the presence of small measurement variations. Therefore, the apparently low reproducibility of DEG detection from current technical replicate tests does not indicate low quality of microarray technology. We also demonstrate that heterogeneous biological variations existing in real cancer data will further reduce the overall reproducibility of DEG detection. Nevertheless, in small subsamples from both simulated and real data, the actual false discovery rate (FDR) for each DEG list tends to be low, suggesting that each separately determined list may comprise mostly true DEGs. Rather than simply counting the overlaps of the discovery lists from different studies for a complex disease, novel metrics are needed for evaluating the reproducibility of discoveries characterized with correlated molecular changes. Supplementaty information: Supplementary data are available at Bioinformatics online.

Versailles Project on Advanced Materials and Standards Interlaboratory Study on Measuring the Thickness and Chemistry of Nanoparticle Coatings Using XPS and LEIS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Belsey, Natalie A.; Cant, David J. H.; Minelli, Caterina

We report the results of a VAMAS (Versailles Project on Advanced Materials and Standards) inter-laboratory study on the measurement of the shell thickness and chemistry of nanoparticle coatings. Peptide-coated gold particles were supplied to laboratories in two forms: a colloidal suspension in pure water and; particles dried onto a silicon wafer. Participants prepared and analyzed these samples using either X-ray photoelectron spectroscopy (XPS) or low energy ion scattering (LEIS). Careful data analysis revealed some significant sources of discrepancy, particularly for XPS. Degradation during transportation, storage or sample preparation resulted in a variability in thickness of 53 %. The calculation methodmore » chosen by XPS participants contributed a variability of 67 %. However, variability of 12 % was achieved for the samples deposited using a single method and by choosing photoelectron peaks that were not adversely affected by instrumental transmission effects. The study identified a need for more consistency in instrumental transmission functions and relative sensitivity factors, since this contributed a variability of 33 %. The results from the LEIS participants were more consistent, with variability of less than 10 % in thickness and this is mostly due to a common method of data analysis. The calculation was performed using a model developed for uniform, flat films and some participants employed a correction factor to account for the sample geometry, which appears warranted based upon a simulation of LEIS data from one of the participants and comparison to the XPS results.« less

Soiling of building envelope surfaces and its effect on solar reflectance – Part III: Interlaboratory study of an accelerated aging method for roofing materials

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sleiman, Mohamad; Chen, Sharon; Gilbert, Haley E.

A laboratory method to simulate natural exposure of roofing materials has been reported in a companion article. Here in the current article, we describe the results of an international, nine-participant interlaboratory study (ILS) conducted in accordance with ASTM Standard E691-09 to establish the precision and reproducibility of this protocol. The accelerated soiling and weathering method was applied four times by each laboratory to replicate coupons of 12 products representing a wide variety of roofing categories (single-ply membrane, factory-applied coating (on metal), bare metal, field-applied coating, asphalt shingle, modified-bitumen cap sheet, clay tile, and concrete tile). Participants reported initial and laboratory-agedmore » values of solar reflectance and thermal emittance. Measured solar reflectances were consistent within and across eight of the nine participating laboratories. Measured thermal emittances reported by six participants exhibited comparable consistency. For solar reflectance, the accelerated aging method is both repeatable and reproducible within an acceptable range of standard deviations: the repeatability standard deviation sr ranged from 0.008 to 0.015 (relative standard deviation of 1.2–2.1%) and the reproducibility standard deviation sR ranged from 0.022 to 0.036 (relative standard deviation of 3.2–5.8%). The ILS confirmed that the accelerated aging method can be reproduced by multiple independent laboratories with acceptable precision. In conclusion, this study supports the adoption of the accelerated aging practice to speed the evaluation and performance rating of new cool roofing materials.« less

Improved porous silicon (P-Si) microarray based PSA (prostate specific antigen) immunoassay by optimized surface density of the capture antibody

PubMed Central

Lee, SangWook; Kim, Soyoun; Malm, Johan; Jeong, Ok Chan; Lilja, Hans; Laurell, Thomas

2014-01-01

Enriching the surface density of immobilized capture antibodies enhances the detection signal of antibody sandwich microarrays. In this study, we improved the detection sensitivity of our previously developed P-Si (porous silicon) antibody microarray by optimizing concentrations of the capturing antibody. We investigated immunoassays using a P-Si microarray at three different capture antibody (PSA - prostate specific antigen) concentrations, analyzing the influence of the antibody density on the assay detection sensitivity. The LOD (limit of detection) for PSA was 2.5ngmL−1, 80pgmL−1, and 800fgmL−1 when arraying the PSA antibody, H117 at the concentration 15µgmL−1, 35µgmL−1 and 154µgmL−1, respectively. We further investigated PSA spiked into human female serum in the range of 800fgmL−1 to 500ngmL−1. The microarray showed a LOD of 800fgmL−1 and a dynamic range of 800 fgmL−1 to 80ngmL−1 in serum spiked samples. PMID:24016590

Comparison of Hepatic 2D Sandwich Cultures and 3D Spheroids for Long-term Toxicity Applications: A Multicenter Study

PubMed Central

Bell, Catherine C; Dankers, Anita C A; Sison-Young, Rowena; Jenkins, Roz; Rowe, Cliff; Goldring, Chris E; Park, Kevin; Regan, Sophie L; Walker, Tracy; Schofield, Chris; Baze, Audrey; Foster, Alison J; Williams, Dominic P; van de Ven, Amy W M; Jacobs, Frank; van Houdt, Jos; Lähteenmäki, Tuula; Snoeys, Jan; Juhila, Satu; Richert, Lysiane; Ingelman-Sundberg, Magnus

2018-01-01

Abstract Primary human hepatocytes (PHHs) are commonly used for in vitro studies of drug-induced liver injury. However, when cultured as 2D monolayers, PHH lose crucial hepatic functions within hours. This dedifferentiation can be ameliorated when PHHs are cultured in sandwich configuration (2Dsw), particularly when cultures are regularly re-overlaid with extracellular matrix, or as 3D spheroids. In this study, the 6 participating laboratories evaluated the robustness of these 2 model systems made from cryopreserved PHH from the same donors considering both inter-donor and inter-laboratory variability and compared their suitability for use in repeated-dose toxicity studies using 5 different hepatotoxins with different toxicity mechanisms. We found that expression levels of proteins involved in drug absorption, distribution, metabolism, and excretion, as well as catalytic activities of 5 different CYPs, were significantly higher in 3D spheroid cultures, potentially affecting the exposure of the cells to drugs and their metabolites. Furthermore, global proteomic analyses revealed that PHH in 3D spheroid configuration were temporally stable whereas proteomes from the same donors in 2Dsw cultures showed substantial alterations in protein expression patterns over the 14 days in culture. Overall, spheroid cultures were more sensitive to the hepatotoxic compounds investigated, particularly upon long-term exposures, across testing sites with little inter-laboratory or inter-donor variability. The data presented here suggest that repeated-dosing regimens improve the predictivity of in vitro toxicity assays, and that PHH spheroids provide a sensitive and robust system for long-term mechanistic studies of drug-induced hepatotoxicity, whereas the 2Dsw system has a more dedifferentiated phenotype and lower sensitivity to detect hepatotoxicity. PMID:29329425

Interlaboratory Study on Differential Analysis of Protein Glycosylation by Mass Spectrometry: The ABRF Glycoprotein Research Multi-Institutional Study 2012*

PubMed Central

Leymarie, Nancy; Griffin, Paula J.; Jonscher, Karen; Kolarich, Daniel; Orlando, Ron; McComb, Mark; Zaia, Joseph; Aguilan, Jennifer; Alley, William R.; Altmann, Friederich; Ball, Lauren E.; Basumallick, Lipika; Bazemore-Walker, Carthene R.; Behnken, Henning; Blank, Michael A.; Brown, Kristy J.; Bunz, Svenja-Catharina; Cairo, Christopher W.; Cipollo, John F.; Daneshfar, Rambod; Desaire, Heather; Drake, Richard R.; Go, Eden P.; Goldman, Radoslav; Gruber, Clemens; Halim, Adnan; Hathout, Yetrib; Hensbergen, Paul J.; Horn, David M.; Hurum, Deanna; Jabs, Wolfgang; Larson, Göran; Ly, Mellisa; Mann, Benjamin F.; Marx, Kristina; Mechref, Yehia; Meyer, Bernd; Möginger, Uwe; Neusüβ, Christian; Nilsson, Jonas; Novotny, Milos V.; Nyalwidhe, Julius O.; Packer, Nicolle H.; Pompach, Petr; Reiz, Bela; Resemann, Anja; Rohrer, Jeffrey S.; Ruthenbeck, Alexandra; Sanda, Miloslav; Schulz, Jan Mirco; Schweiger-Hufnagel, Ulrike; Sihlbom, Carina; Song, Ehwang; Staples, Gregory O.; Suckau, Detlev; Tang, Haixu; Thaysen-Andersen, Morten; Viner, Rosa I.; An, Yanming; Valmu, Leena; Wada, Yoshinao; Watson, Megan; Windwarder, Markus; Whittal, Randy; Wuhrer, Manfred; Zhu, Yiying; Zou, Chunxia

2013-01-01

One of the principal goals of glycoprotein research is to correlate glycan structure and function. Such correlation is necessary in order for one to understand the mechanisms whereby glycoprotein structure elaborates the functions of myriad proteins. The accurate comparison of glycoforms and quantification of glycosites are essential steps in this direction. Mass spectrometry has emerged as a powerful analytical technique in the field of glycoprotein characterization. Its sensitivity, high dynamic range, and mass accuracy provide both quantitative and sequence/structural information. As part of the 2012 ABRF Glycoprotein Research Group study, we explored the use of mass spectrometry and ancillary methodologies to characterize the glycoforms of two sources of human prostate specific antigen (PSA). PSA is used as a tumor marker for prostate cancer, with increasing blood levels used to distinguish between normal and cancer states. The glycans on PSA are believed to be biantennary N-linked, and it has been observed that prostate cancer tissues and cell lines contain more antennae than their benign counterparts. Thus, the ability to quantify differences in glycosylation associated with cancer has the potential to positively impact the use of PSA as a biomarker. We studied standard peptide-based proteomics/glycomics methodologies, including LC-MS/MS for peptide/glycopeptide sequencing and label-free approaches for differential quantification. We performed an interlaboratory study to determine the ability of different laboratories to correctly characterize the differences between glycoforms from two different sources using mass spectrometry methods. We used clustering analysis and ancillary statistical data treatment on the data sets submitted by participating laboratories to obtain a consensus of the glycoforms and abundances. The results demonstrate the relative strengths and weaknesses of top-down glycoproteomics, bottom-up glycoproteomics, and glycomics methods. PMID:23764502

Identification of candidate genes in osteoporosis by integrated microarray analysis.

PubMed

Li, J J; Wang, B Q; Fei, Q; Yang, Y; Li, D

2016-12-01

In order to screen the altered gene expression profile in peripheral blood mononuclear cells of patients with osteoporosis, we performed an integrated analysis of the online microarray studies of osteoporosis. We searched the Gene Expression Omnibus (GEO) database for microarray studies of peripheral blood mononuclear cells in patients with osteoporosis. Subsequently, we integrated gene expression data sets from multiple microarray studies to obtain differentially expressed genes (DEGs) between patients with osteoporosis and normal controls. Gene function analysis was performed to uncover the functions of identified DEGs. A total of three microarray studies were selected for integrated analysis. In all, 1125 genes were found to be significantly differentially expressed between osteoporosis patients and normal controls, with 373 upregulated and 752 downregulated genes. Positive regulation of the cellular amino metabolic process (gene ontology (GO): 0033240, false discovery rate (FDR) = 1.00E + 00) was significantly enriched under the GO category for biological processes, while for molecular functions, flavin adenine dinucleotide binding (GO: 0050660, FDR = 3.66E-01) and androgen receptor binding (GO: 0050681, FDR = 6.35E-01) were significantly enriched. DEGs were enriched in many osteoporosis-related signalling pathways, including those of mitogen-activated protein kinase (MAPK) and calcium. Protein-protein interaction (PPI) network analysis showed that the significant hub proteins contained ubiquitin specific peptidase 9, X-linked (Degree = 99), ubiquitin specific peptidase 19 (Degree = 57) and ubiquitin conjugating enzyme E2 B (Degree = 57). Analysis of gene function of identified differentially expressed genes may expand our understanding of fundamental mechanisms leading to osteoporosis. Moreover, significantly enriched pathways, such as MAPK and calcium, may involve in osteoporosis through osteoblastic differentiation and bone formation.Cite this article: J. J. Li, B. Q. Wang, Q. Fei, Y. Yang, D. Li. Identification of candidate genes in osteoporosis by integrated microarray analysis. Bone Joint Res 2016;5:594-601. DOI: 10.1302/2046-3758.512.BJR-2016-0073.R1. © 2016 Fei et al.

cDNA microarray analysis of esophageal cancer: discoveries and prospects.

PubMed

Shimada, Yutaka; Sato, Fumiaki; Shimizu, Kazuharu; Tsujimoto, Gozoh; Tsukada, Kazuhiro

2009-07-01

Recent progress in molecular biology has revealed many genetic and epigenetic alterations that are involved in the development and progression of esophageal cancer. Microarray analysis has also revealed several genetic networks that are involved in esophageal cancer. However, clinical application of microarray techniques and use of microarray data have not yet occurred. In this review, we focus on the recent developments and problems with microarray analysis of esophageal cancer.

Manufacturing of microarrays.

PubMed

Petersen, David W; Kawasaki, Ernest S

2007-01-01

DNA microarray technology has become a powerful tool in the arsenal of the molecular biologist. Capitalizing on high precision robotics and the wealth of DNA sequences annotated from the genomes of a large number of organisms, the manufacture of microarrays is now possible for the average academic laboratory with the funds and motivation. Microarray production requires attention to both biological and physical resources, including DNA libraries, robotics, and qualified personnel. While the fabrication of microarrays is a very labor-intensive process, production of quality microarrays individually tailored on a project-by-project basis will help researchers shed light on future scientific questions.

The Longhorn Array Database (LAD): An Open-Source, MIAME compliant implementation of the Stanford Microarray Database (SMD)

PubMed Central

Killion, Patrick J; Sherlock, Gavin; Iyer, Vishwanath R

2003-01-01

Background The power of microarray analysis can be realized only if data is systematically archived and linked to biological annotations as well as analysis algorithms. Description The Longhorn Array Database (LAD) is a MIAME compliant microarray database that operates on PostgreSQL and Linux. It is a fully open source version of the Stanford Microarray Database (SMD), one of the largest microarray databases. LAD is available at Conclusions Our development of LAD provides a simple, free, open, reliable and proven solution for storage and analysis of two-color microarray data. PMID:12930545

The emergence and diffusion of DNA microarray technology.

PubMed

Lenoir, Tim; Giannella, Eric

2006-08-22

The network model of innovation widely adopted among researchers in the economics of science and technology posits relatively porous boundaries between firms and academic research programs and a bi-directional flow of inventions, personnel, and tacit knowledge between sites of university and industry innovation. Moreover, the model suggests that these bi-directional flows should be considered as mutual stimulation of research and invention in both industry and academe, operating as a positive feedback loop. One side of this bi-directional flow--namely; the flow of inventions into industry through the licensing of university-based technologies--has been well studied; but the reverse phenomenon of the stimulation of university research through the absorption of new directions emanating from industry has yet to be investigated in much detail. We discuss the role of federal funding of academic research in the microarray field, and the multiple pathways through which federally supported development of commercial microarray technologies have transformed core academic research fields. Our study confirms the picture put forward by several scholars that the open character of networked economies is what makes them truly innovative. In an open system innovations emerge from the network. The emergence and diffusion of microarray technologies we have traced here provides an excellent example of an open system of innovation in action. Whether they originated in a startup company environment that operated like a think-tank, such as Affymax, the research labs of a large firm, such as Agilent, or within a research university, the inventors we have followed drew heavily on knowledge resources from all parts of the network in bringing microarray platforms to light. Federal funding for high-tech startups and new industrial development was important at several phases in the early history of microarrays, and federal funding of academic researchers using microarrays was fundamental to transforming the research agendas of several fields within academe. The typical story told about the role of federal funding emphasizes the spillovers from federally funded academic research to industry. Our study shows that the knowledge spillovers worked both ways, with federal funding of non-university research providing the impetus for reshaping the research agendas of several academic fields.

The emergence and diffusion of DNA microarray technology

PubMed Central

Lenoir, Tim; Giannella, Eric

2006-01-01

The network model of innovation widely adopted among researchers in the economics of science and technology posits relatively porous boundaries between firms and academic research programs and a bi-directional flow of inventions, personnel, and tacit knowledge between sites of university and industry innovation. Moreover, the model suggests that these bi-directional flows should be considered as mutual stimulation of research and invention in both industry and academe, operating as a positive feedback loop. One side of this bi-directional flow – namely; the flow of inventions into industry through the licensing of university-based technologies – has been well studied; but the reverse phenomenon of the stimulation of university research through the absorption of new directions emanating from industry has yet to be investigated in much detail. We discuss the role of federal funding of academic research in the microarray field, and the multiple pathways through which federally supported development of commercial microarray technologies have transformed core academic research fields. Our study confirms the picture put forward by several scholars that the open character of networked economies is what makes them truly innovative. In an open system innovations emerge from the network. The emergence and diffusion of microarray technologies we have traced here provides an excellent example of an open system of innovation in action. Whether they originated in a startup company environment that operated like a think-tank, such as Affymax, the research labs of a large firm, such as Agilent, or within a research university, the inventors we have followed drew heavily on knowledge resources from all parts of the network in bringing microarray platforms to light. Federal funding for high-tech startups and new industrial development was important at several phases in the early history of microarrays, and federal funding of academic researchers using microarrays was fundamental to transforming the research agendas of several fields within academe. The typical story told about the role of federal funding emphasizes the spillovers from federally funded academic research to industry. Our study shows that the knowledge spillovers worked both ways, with federal funding of non-university research providing the impetus for reshaping the research agendas of several academic fields. PMID:16925816

IDENTIFICATION OF BIOLOGICALLY RELEVANT GENES USING A DATABASE OF RAT LIVER AND KIDNEY BASELINE GENE EXPRESSION

EPA Science Inventory

Microarray data from independent labs and studies can be compared to potentially identify toxicologically and biologically relevant genes. The Baseline Animal Database working group of HESI was formed to assess baseline gene expression from microarray data derived from control or...

Transcriptome analysis of salinity stress responses in common wheat using a 22k oligo-DNA microarray.

PubMed

Kawaura, Kanako; Mochida, Keiichi; Yamazaki, Yukiko; Ogihara, Yasunari

2006-04-01

In this study, we constructed a 22k wheat oligo-DNA microarray. A total of 148,676 expressed sequence tags of common wheat were collected from the database of the Wheat Genomics Consortium of Japan. These were grouped into 34,064 contigs, which were then used to design an oligonucleotide DNA microarray. Following a multistep selection of the sense strand, 21,939 60-mer oligo-DNA probes were selected for attachment on the microarray slide. This 22k oligo-DNA microarray was used to examine the transcriptional response of wheat to salt stress. More than 95% of the probes gave reproducible hybridization signals when targeted with RNAs extracted from salt-treated wheat shoots and roots. With the microarray, we identified 1,811 genes whose expressions changed more than 2-fold in response to salt. These included genes known to mediate response to salt, as well as unknown genes, and they were classified into 12 major groups by hierarchical clustering. These gene expression patterns were also confirmed by real-time reverse transcription-PCR. Many of the genes with unknown function were clustered together with genes known to be involved in response to salt stress. Thus, analysis of gene expression patterns combined with gene ontology should help identify the function of the unknown genes. Also, functional analysis of these wheat genes should provide new insight into the response to salt stress. Finally, these results indicate that the 22k oligo-DNA microarray is a reliable method for monitoring global gene expression patterns in wheat.

MADGE: scalable distributed data management software for cDNA microarrays.

PubMed

McIndoe, Richard A; Lanzen, Aaron; Hurtz, Kimberly

2003-01-01

The human genome project and the development of new high-throughput technologies have created unparalleled opportunities to study the mechanism of diseases, monitor the disease progression and evaluate effective therapies. Gene expression profiling is a critical tool to accomplish these goals. The use of nucleic acid microarrays to assess the gene expression of thousands of genes simultaneously has seen phenomenal growth over the past five years. Although commercial sources of microarrays exist, investigators wanting more flexibility in the genes represented on the array will turn to in-house production. The creation and use of cDNA microarrays is a complicated process that generates an enormous amount of information. Effective data management of this information is essential to efficiently access, analyze, troubleshoot and evaluate the microarray experiments. We have developed a distributable software package designed to track and store the various pieces of data generated by a cDNA microarray facility. This includes the clone collection storage data, annotation data, workflow queues, microarray data, data repositories, sample submission information, and project/investigator information. This application was designed using a 3-tier client server model. The data access layer (1st tier) contains the relational database system tuned to support a large number of transactions. The data services layer (2nd tier) is a distributed COM server with full database transaction support. The application layer (3rd tier) is an internet based user interface that contains both client and server side code for dynamic interactions with the user. This software is freely available to academic institutions and non-profit organizations at http://www.genomics.mcg.edu/niddkbtc.

Intra- and inter-laboratory reliability of a cryopreserved trout hepatocyte assay for the prediction of chemical bioaccumulation potential

EPA Science Inventory

Cryopreserved trout hepatocytes provide a convenient in vitro system for measuring the intrinsic clearance of xenobiotics. Measured clearance rates can then be extrapolated to the whole animal as a means of improving modeled bioaccumulation predictions. To date, however, the in...

Evaluation of Methods for Analysis of Lead in Air Particulates: An Intra-Laboratory and Inter-Laboratory Comparison

EPA Science Inventory

In 2008, the United States Environmental Protection Agency (USEPA) set a new National Ambient Air Quality Standard (NAAQS) for lead (Pb) in total suspended particulate matter (Pb-TSP) which called for significant decreases in the allowable limits. The Federal Reference Method (FR...

Evaluation of the repeatability and reproducibility of a suite of qPCR based microbial source tracking methods

EPA Science Inventory

Many PCR-based methods for microbial source tracking (MST) have been developed and validated within individual research laboratories. Inter-laboratory validation of these methods, however, has been minimal, and the effects of protocol standardization regimes have not been thor...

Inter-laboratory Comparison of Real-time PCR Methods for Quantification of General Fecal Indicator Bacteria

EPA Science Inventory

The application of quantitative real-time PCR (qPCR) technologies for the rapid identification of fecal bacteria in environmental waters is being considered for use as a national water quality metric in the United States. The transition from research tool to a standardized prot...

Interlaboratory Comparison of Real-time PCR Protocols for Quantification of General Fecal Indicator Bacteria

EPA Science Inventory

The application of quantitative real-time PCR (qPCR) technologies for the rapid identification of fecal bacteria in environmental waters is being considered for use as a national water quality metric in the United States. The transition from research tool to a standardized proto...

In vitro to In vivo extrapolation of hepatic metabolism in fish: An inter-laboratory comparison of In vitro methods

EPA Science Inventory

Chemical biotransformation represents the single largest source of uncertainty in chemical bioaccumulation assessments for fish. In vitro methods employing isolated hepatocytes and liver subcellular fractions (S9) can be used to estimate whole-body rates of chemical metabolism, ...

Touryan Elected Chairman

Science.gov Websites

Touryan Elected Chairman of Proliferation Prevention Advisory Board For information contact: e:mail ) National Renewable Energy Laboratory (NREL) was elected chairman of the Inter-Laboratory Advisory Board for opportunities and a means of rapid commercialization of defense technologies. It creates a win-win situation for

INTERLABORATORY VALIDATION OF USEPA METHOD 1680: FECAL COLIFORMS IN BIOSOLIDS BY MULTIPLE-TUBE FERMENTATION PROCEDURES

EPA Science Inventory

In the US, the use and disposal of biosolids (including domestic septage) are regulated under 40 CFR Part 503. Subpart D of this regulation protects public health and the environment through requirements designed to reduce the potential for contact with pathogens in biosolids app...

INTERLABORATORY VALIDATION OF USEPA METHOD 1680: FECAL COLIFORMS IN BIOSOLIDS BY MULTIPLE-TUBE FERMENTATION PROCEDURES

EPA Science Inventory

In the United States, the use and disposal of biosolids (including domestic septage) are regulated under 40 CFR Part 503. Subpart D of this regulation protects public health and the environment through requirements designed to reduce the potential for contact with pathogens in bi...

Analysis of Protein-DNA Interaction by Chromatin Immunoprecipitation and DNA Tiling Microarray (ChIP-on-chip).

PubMed

Gao, Hui; Zhao, Chunyan

2018-01-01

Chromatin immunoprecipitation (ChIP) has become the most effective and widely used tool to study the interactions between specific proteins or modified forms of proteins and a genomic DNA region. Combined with genome-wide profiling technologies, such as microarray hybridization (ChIP-on-chip) or massively parallel sequencing (ChIP-seq), ChIP could provide a genome-wide mapping of in vivo protein-DNA interactions in various organisms. Here, we describe a protocol of ChIP-on-chip that uses tiling microarray to obtain a genome-wide profiling of ChIPed DNA.

Interlaboratory comparability, bias, and precision for four laboratories measuring constituents in precipitation, November 1982-August 1983

USGS Publications Warehouse

Brooks, M.H.; Schroder, L.J.; Malo, B.A.

1985-01-01

Four laboratories were evaluated in their analysis of identical natural and simulated precipitation water samples. Interlaboratory comparability was evaluated using analysis of variance coupled with Duncan 's multiple range test, and linear-regression models describing the relations between individual laboratory analytical results for natural precipitation samples. Results of the statistical analyses indicate that certain pairs of laboratories produce different results when analyzing identical samples. Analyte bias for each laboratory was examined using analysis of variance coupled with Duncan 's multiple range test on data produced by the laboratories from the analysis of identical simulated precipitation samples. Bias for a given analyte produced by a single laboratory has been indicated when the laboratory mean for that analyte is shown to be significantly different from the mean for the most-probable analyte concentrations in the simulated precipitation samples. Ion-chromatographic methods for the determination of chloride, nitrate, and sulfate have been compared with the colorimetric methods that were also in use during the study period. Comparisons were made using analysis of variance coupled with Duncan 's multiple range test for means produced by the two methods. Analyte precision for each laboratory has been estimated by calculating a pooled variance for each analyte. Analyte estimated precisions have been compared using F-tests and differences in analyte precisions for laboratory pairs have been reported. (USGS)

EzArray: A web-based highly automated Affymetrix expression array data management and analysis system

PubMed Central

Zhu, Yuerong; Zhu, Yuelin; Xu, Wei

2008-01-01

Background Though microarray experiments are very popular in life science research, managing and analyzing microarray data are still challenging tasks for many biologists. Most microarray programs require users to have sophisticated knowledge of mathematics, statistics and computer skills for usage. With accumulating microarray data deposited in public databases, easy-to-use programs to re-analyze previously published microarray data are in high demand. Results EzArray is a web-based Affymetrix expression array data management and analysis system for researchers who need to organize microarray data efficiently and get data analyzed instantly. EzArray organizes microarray data into projects that can be analyzed online with predefined or custom procedures. EzArray performs data preprocessing and detection of differentially expressed genes with statistical methods. All analysis procedures are optimized and highly automated so that even novice users with limited pre-knowledge of microarray data analysis can complete initial analysis quickly. Since all input files, analysis parameters, and executed scripts can be downloaded, EzArray provides maximum reproducibility for each analysis. In addition, EzArray integrates with Gene Expression Omnibus (GEO) and allows instantaneous re-analysis of published array data. Conclusion EzArray is a novel Affymetrix expression array data analysis and sharing system. EzArray provides easy-to-use tools for re-analyzing published microarray data and will help both novice and experienced users perform initial analysis of their microarray data from the location of data storage. We believe EzArray will be a useful system for facilities with microarray services and laboratories with multiple members involved in microarray data analysis. EzArray is freely available from . PMID:18218103

A Java-based tool for the design of classification microarrays.

PubMed

Meng, Da; Broschat, Shira L; Call, Douglas R

2008-08-04

Classification microarrays are used for purposes such as identifying strains of bacteria and determining genetic relationships to understand the epidemiology of an infectious disease. For these cases, mixed microarrays, which are composed of DNA from more than one organism, are more effective than conventional microarrays composed of DNA from a single organism. Selection of probes is a key factor in designing successful mixed microarrays because redundant sequences are inefficient and limited representation of diversity can restrict application of the microarray. We have developed a Java-based software tool, called PLASMID, for use in selecting the minimum set of probe sequences needed to classify different groups of plasmids or bacteria. The software program was successfully applied to several different sets of data. The utility of PLASMID was illustrated using existing mixed-plasmid microarray data as well as data from a virtual mixed-genome microarray constructed from different strains of Streptococcus. Moreover, use of data from expression microarray experiments demonstrated the generality of PLASMID. In this paper we describe a new software tool for selecting a set of probes for a classification microarray. While the tool was developed for the design of mixed microarrays-and mixed-plasmid microarrays in particular-it can also be used to design expression arrays. The user can choose from several clustering methods (including hierarchical, non-hierarchical, and a model-based genetic algorithm), several probe ranking methods, and several different display methods. A novel approach is used for probe redundancy reduction, and probe selection is accomplished via stepwise discriminant analysis. Data can be entered in different formats (including Excel and comma-delimited text), and dendrogram, heat map, and scatter plot images can be saved in several different formats (including jpeg and tiff). Weights generated using stepwise discriminant analysis can be stored for analysis of subsequent experimental data. Additionally, PLASMID can be used to construct virtual microarrays with genomes from public databases, which can then be used to identify an optimal set of probes.

Evaluating reproducibility of differential expression discoveries in microarray studies by considering correlated molecular changes.

PubMed

Zhang, Min; Zhang, Lin; Zou, Jinfeng; Yao, Chen; Xiao, Hui; Liu, Qing; Wang, Jing; Wang, Dong; Wang, Chenguang; Guo, Zheng

2009-07-01

According to current consistency metrics such as percentage of overlapping genes (POG), lists of differentially expressed genes (DEGs) detected from different microarray studies for a complex disease are often highly inconsistent. This irreproducibility problem also exists in other high-throughput post-genomic areas such as proteomics and metabolism. A complex disease is often characterized with many coordinated molecular changes, which should be considered when evaluating the reproducibility of discovery lists from different studies. We proposed metrics percentage of overlapping genes-related (POGR) and normalized POGR (nPOGR) to evaluate the consistency between two DEG lists for a complex disease, considering correlated molecular changes rather than only counting gene overlaps between the lists. Based on microarray datasets of three diseases, we showed that though the POG scores for DEG lists from different studies for each disease are extremely low, the POGR and nPOGR scores can be rather high, suggesting that the apparently inconsistent DEG lists may be highly reproducible in the sense that they are actually significantly correlated. Observing different discovery results for a disease by the POGR and nPOGR scores will obviously reduce the uncertainty of the microarray studies. The proposed metrics could also be applicable in many other high-throughput post-genomic areas.

THE ABRF MARG MICROARRAY SURVEY 2005: TAKING THE PULSE ON THE MICROARRAY FIELD

EPA Science Inventory

Over the past several years microarray technology has evolved into a critical component of any discovery based program. Since 1999, the Association of Biomolecular Resource Facilities (ABRF) Microarray Research Group (MARG) has conducted biennial surveys designed to generate a pr...

Development of a Digital Microarray with Interferometric Reflectance Imaging

NASA Astrophysics Data System (ADS)

Sevenler, Derin

This dissertation describes a new type of molecular assay for nucleic acids and proteins. We call this technique a digital microarray since it is conceptually similar to conventional fluorescence microarrays, yet it performs enumerative ('digital') counting of the number captured molecules. Digital microarrays are approximately 10,000-fold more sensitive than fluorescence microarrays, yet maintain all of the strengths of the platform including low cost and high multiplexing (i.e., many different tests on the same sample simultaneously). Digital microarrays use gold nanorods to label the captured target molecules. Each gold nanorod on the array is individually detected based on its light scattering, with an interferometric microscopy technique called SP-IRIS. Our optimized high-throughput version of SP-IRIS is able to scan a typical array of 500 spots in less than 10 minutes. Digital DNA microarrays may have utility in applications where sequencing is prohibitively expensive or slow. As an example, we describe a digital microarray assay for gene expression markers of bacterial drug resistance.

Implementation of mutual information and bayes theorem for classification microarray data

NASA Astrophysics Data System (ADS)

Dwifebri Purbolaksono, Mahendra; Widiastuti, Kurnia C.; Syahrul Mubarok, Mohamad; Adiwijaya; Aminy Ma’ruf, Firda

2018-03-01

Microarray Technology is one of technology which able to read the structure of gen. The analysis is important for this technology. It is for deciding which attribute is more important than the others. Microarray technology is able to get cancer information to diagnose a person’s gen. Preparation of microarray data is a huge problem and takes a long time. That is because microarray data contains high number of insignificant and irrelevant attributes. So, it needs a method to reduce the dimension of microarray data without eliminating important information in every attribute. This research uses Mutual Information to reduce dimension. System is built with Machine Learning approach specifically Bayes Theorem. This theorem uses a statistical and probability approach. By combining both methods, it will be powerful for Microarray Data Classification. The experiment results show that system is good to classify Microarray data with highest F1-score using Bayesian Network by 91.06%, and Naïve Bayes by 88.85%.

Protein microarray with horseradish peroxidase chemiluminescence for quantification of serum α-fetoprotein.

PubMed

Zhao, Yuanshun; Zhang, Yonghong; Lin, Dongdong; Li, Kang; Yin, Chengzeng; Liu, Xiuhong; Jin, Boxun; Sun, Libo; Liu, Jinhua; Zhang, Aiying; Li, Ning

2015-10-01

To develop and evaluate a protein microarray assay with horseradish peroxidase (HRP) chemiluminescence for quantification of α-fetoprotein (AFP) in serum from patients with hepatocellular carcinoma (HCC). A protein microarray assay for AFP was developed. Serum was collected from patients with HCC and healthy control subjects. AFP was quantified using protein microarray and enzyme-linked immunosorbent assay (ELISA). Serum AFP concentrations determined via protein microarray were positively correlated (r = 0.973) with those determined via ELISA in patients with HCC (n = 60) and healthy control subjects (n = 30). Protein microarray showed 80% sensitivity and 100% specificity for HCC diagnosis. ELISA had 83.3% sensitivity and 100% specificity. Protein microarray effectively distinguished between patients with HCC and healthy control subjects (area under ROC curve 0.974; 95% CI 0.000, 1.000). Protein microarray is a rapid, simple and low-cost alternative to ELISA for detecting AFP in human serum. © The Author(s) 2015.

A multiplex reverse transcription PCR and automated electronic microarray assay for detection and differentiation of seven viruses affecting swine.

PubMed

Erickson, A; Fisher, M; Furukawa-Stoffer, T; Ambagala, A; Hodko, D; Pasick, J; King, D P; Nfon, C; Ortega Polo, R; Lung, O

2018-04-01

Microarray technology can be useful for pathogen detection as it allows simultaneous interrogation of the presence or absence of a large number of genetic signatures. However, most microarray assays are labour-intensive and time-consuming to perform. This study describes the development and initial evaluation of a multiplex reverse transcription (RT)-PCR and novel accompanying automated electronic microarray assay for simultaneous detection and differentiation of seven important viruses that affect swine (foot-and-mouth disease virus [FMDV], swine vesicular disease virus [SVDV], vesicular exanthema of swine virus [VESV], African swine fever virus [ASFV], classical swine fever virus [CSFV], porcine respiratory and reproductive syndrome virus [PRRSV] and porcine circovirus type 2 [PCV2]). The novel electronic microarray assay utilizes a single, user-friendly instrument that integrates and automates capture probe printing, hybridization, washing and reporting on a disposable electronic microarray cartridge with 400 features. This assay accurately detected and identified a total of 68 isolates of the seven targeted virus species including 23 samples of FMDV, representing all seven serotypes, and 10 CSFV strains, representing all three genotypes. The assay successfully detected viruses in clinical samples from the field, experimentally infected animals (as early as 1 day post-infection (dpi) for FMDV and SVDV, 4 dpi for ASFV, 5 dpi for CSFV), as well as in biological material that were spiked with target viruses. The limit of detection was 10 copies/μl for ASFV, PCV2 and PRRSV, 100 copies/μl for SVDV, CSFV, VESV and 1,000 copies/μl for FMDV. The electronic microarray component had reduced analytical sensitivity for several of the target viruses when compared with the multiplex RT-PCR. The integration of capture probe printing allows custom onsite array printing as needed, while electrophoretically driven hybridization generates results faster than conventional microarrays that rely on passive hybridization. With further refinement, this novel, rapid, highly automated microarray technology has potential applications in multipathogen surveillance of livestock diseases. © 2017 Her Majesty the Queen in Right of Canada • Transboundary and Emerging Diseases.

Microarray analysis of potential genes in the pathogenesis of recurrent oral ulcer.

PubMed

Han, Jingying; He, Zhiwei; Li, Kun; Hou, Lu

2015-01-01

Recurrent oral ulcer seriously threatens patients' daily life and health. This study investigated potential genes and pathways that participate in the pathogenesis of recurrent oral ulcer by high throughput bioinformatic analysis. RT-PCR and Western blot were applied to further verify screened interleukins effect. Recurrent oral ulcer related genes were collected from websites and papers, and further found out from Human Genome 280 6.0 microarray data. Each pathway of recurrent oral ulcer related genes were got through chip hybridization. RT-PCR was applied to test four recurrent oral ulcer related genes to verify the microarray data. Data transformation, scatter plot, clustering analysis, and expression pattern analysis were used to analyze recurrent oral ulcer related gene expression changes. Recurrent oral ulcer gene microarray was successfully established. Microarray showed that 551 genes involved in recurrent oral ulcer activity and 196 genes were recurrent oral ulcer related genes. Of them, 76 genes up-regulated, 62 genes down-regulated, and 58 genes up-/down-regulated. Total expression level up-regulated 752 times (60%) and down-regulated 485 times (40%). IL-2 plays an important role in the occurrence, development and recurrence of recurrent oral ulcer on the mRNA and protein levels. Gene microarray can be used to analyze potential genes and pathways in recurrent oral ulcer. IL-2 may be involved in the pathogenesis of recurrent oral ulcer.

A proposed metric for assessing the measurement quality of individual microarrays

PubMed Central

Kim, Kyoungmi; Page, Grier P; Beasley, T Mark; Barnes, Stephen; Scheirer, Katherine E; Allison, David B

2006-01-01

Background High-density microarray technology is increasingly applied to study gene expression levels on a large scale. Microarray experiments rely on several critical steps that may introduce error and uncertainty in analyses. These steps include mRNA sample extraction, amplification and labeling, hybridization, and scanning. In some cases this may be manifested as systematic spatial variation on the surface of microarray in which expression measurements within an individual array may vary as a function of geographic position on the array surface. Results We hypothesized that an index of the degree of spatiality of gene expression measurements associated with their physical geographic locations on an array could indicate the summary of the physical reliability of the microarray. We introduced a novel way to formulate this index using a statistical analysis tool. Our approach regressed gene expression intensity measurements on a polynomial response surface of the microarray's Cartesian coordinates. We demonstrated this method using a fixed model and presented results from real and simulated datasets. Conclusion We demonstrated the potential of such a quantitative metric for assessing the reliability of individual arrays. Moreover, we showed that this procedure can be incorporated into laboratory practice as a means to set quality control specifications and as a tool to determine whether an array has sufficient quality to be retained in terms of spatial correlation of gene expression measurements. PMID:16430768

Intra-/inter-laboratory validation study on reactive oxygen species assay for chemical photosafety evaluation using two different solar simulators.

PubMed

Onoue, Satomi; Hosoi, Kazuhiro; Toda, Tsuguto; Takagi, Hironori; Osaki, Naoto; Matsumoto, Yasuhiro; Kawakami, Satoru; Wakuri, Shinobu; Iwase, Yumiko; Yamamoto, Toshinobu; Nakamura, Kazuichi; Ohno, Yasuo; Kojima, Hajime

2014-06-01

A previous multi-center validation study demonstrated high transferability and reliability of reactive oxygen species (ROS) assay for photosafety evaluation. The present validation study was undertaken to verify further the applicability of different solar simulators and assay performance. In 7 participating laboratories, 2 standards and 42 coded chemicals, including 23 phototoxins and 19 non-phototoxic drugs/chemicals, were assessed by the ROS assay using two different solar simulators (Atlas Suntest CPS series, 3 labs; and Seric SXL-2500V2, 4 labs). Irradiation conditions could be optimized using quinine and sulisobenzone as positive and negative standards to offer consistent assay outcomes. In both solar simulators, the intra- and inter-day precisions (coefficient of variation; CV) for quinine were found to be below 10%. The inter-laboratory CV for quinine averaged 15.4% (Atlas Suntest CPS) and 13.2% (Seric SXL-2500V2) for singlet oxygen and 17.0% (Atlas Suntest CPS) and 7.1% (Seric SXL-2500V2) for superoxide, suggesting high inter-laboratory reproducibility even though different solar simulators were employed for the ROS assay. In the ROS assay on 42 coded chemicals, some chemicals (ca. 19-29%) were unevaluable because of limited solubility and spectral interference. Although several false positives appeared with positive predictivity of ca. 76-92% (Atlas Suntest CPS) and ca. 75-84% (Seric SXL-2500V2), there were no false negative predictions in both solar simulators. A multi-center validation study on the ROS assay demonstrated satisfactory transferability, accuracy, precision, and predictivity, as well as the availability of other solar simulators. Copyright © 2013 Elsevier Ltd. All rights reserved.

Microfluidic extraction and microarray detection of biomarkers from cancer tissue slides

NASA Astrophysics Data System (ADS)

Nguyen, H. T.; Dupont, L. N.; Jean, A. M.; Géhin, T.; Chevolot, Y.; Laurenceau, E.; Gijs, M. A. M.

2018-03-01

We report here a new microfluidic method allowing for the quantification of human epidermal growth factor receptor 2 (HER2) expression levels from formalin-fixed breast cancer tissues. After partial extraction of proteins from the tissue slide, the extract is routed to an antibody (Ab) microarray for HER2 titration by fluorescence. Then the HER2-expressing cell area is evaluated by immunofluorescence (IF) staining of the tissue slide and used to normalize the fluorescent HER2 signal measured from the Ab microarray. The number of HER2 gene copies measured by fluorescence in situ hybridization (FISH) on an adjacent tissue slide is concordant with the normalized HER2 expression signal. This work is the first study implementing biomarker extraction and detection from cancer tissue slides using microfluidics in combination with a microarray system, paving the way for further developments towards multiplex and precise quantification of cancer biomarkers.

Glycan microarray screening assay for glycosyltransferase specificities.

PubMed

Peng, Wenjie; Nycholat, Corwin M; Razi, Nahid

2013-01-01

Glycan microarrays represent a high-throughput approach to determining the specificity of glycan-binding proteins against a large set of glycans in a single format. This chapter describes the use of a glycan microarray platform for evaluating the activity and substrate specificity of glycosyltransferases (GTs). The methodology allows simultaneous screening of hundreds of immobilized glycan acceptor substrates by in situ incubation of a GT and its appropriate donor substrate on the microarray surface. Using biotin-conjugated donor substrate enables direct detection of the incorporated sugar residues on acceptor substrates on the array. In addition, the feasibility of the method has been validated using label-free donor substrate combined with lectin-based detection of product to assess enzyme activity. Here, we describe the application of both procedures to assess the specificity of a recombinant human α2-6 sialyltransferase. This technique is readily adaptable to studying other glycosyltransferases.

LS-CAP: an algorithm for identifying cytogenetic aberrations in hepatocellular carcinoma using microarray data.

PubMed

He, Xianmin; Wei, Qing; Sun, Meiqian; Fu, Xuping; Fan, Sichang; Li, Yao

2006-05-01

Biological techniques such as Array-Comparative genomic hybridization (CGH), fluorescent in situ hybridization (FISH) and affymetrix single nucleotide pleomorphism (SNP) array have been used to detect cytogenetic aberrations. However, on genomic scale, these techniques are labor intensive and time consuming. Comparative genomic microarray analysis (CGMA) has been used to identify cytogenetic changes in hepatocellular carcinoma (HCC) using gene expression microarray data. However, CGMA algorithm can not give precise localization of aberrations, fails to identify small cytogenetic changes, and exhibits false negatives and positives. Locally un-weighted smoothing cytogenetic aberrations prediction (LS-CAP) based on local smoothing and binomial distribution can be expected to address these problems. LS-CAP algorithm was built and used on HCC microarray profiles. Eighteen cytogenetic abnormalities were identified, among them 5 were reported previously, and 12 were proven by CGH studies. LS-CAP effectively reduced the false negatives and positives, and precisely located small fragments with cytogenetic aberrations.

MAPPI-DAT: data management and analysis for protein-protein interaction data from the high-throughput MAPPIT cell microarray platform.

PubMed

Gupta, Surya; De Puysseleyr, Veronic; Van der Heyden, José; Maddelein, Davy; Lemmens, Irma; Lievens, Sam; Degroeve, Sven; Tavernier, Jan; Martens, Lennart

2017-05-01

Protein-protein interaction (PPI) studies have dramatically expanded our knowledge about cellular behaviour and development in different conditions. A multitude of high-throughput PPI techniques have been developed to achieve proteome-scale coverage for PPI studies, including the microarray based Mammalian Protein-Protein Interaction Trap (MAPPIT) system. Because such high-throughput techniques typically report thousands of interactions, managing and analysing the large amounts of acquired data is a challenge. We have therefore built the MAPPIT cell microArray Protein Protein Interaction-Data management & Analysis Tool (MAPPI-DAT) as an automated data management and analysis tool for MAPPIT cell microarray experiments. MAPPI-DAT stores the experimental data and metadata in a systematic and structured way, automates data analysis and interpretation, and enables the meta-analysis of MAPPIT cell microarray data across all stored experiments. MAPPI-DAT is developed in Python, using R for data analysis and MySQL as data management system. MAPPI-DAT is cross-platform and can be ran on Microsoft Windows, Linux and OS X/macOS. The source code and a Microsoft Windows executable are freely available under the permissive Apache2 open source license at https://github.com/compomics/MAPPI-DAT. jan.tavernier@vib-ugent.be or lennart.martens@vib-ugent.be. Supplementary data are available at Bioinformatics online. © The Author 2017. Published by Oxford University Press.

Statistical issues in signal extraction from microarrays

NASA Astrophysics Data System (ADS)

Bergemann, Tracy; Quiaoit, Filemon; Delrow, Jeffrey J.; Zhao, Lue Ping

2001-06-01

Microarray technologies are increasingly used in biomedical research to study genome-wide expression profiles in the post genomic era. Their popularity is largely due to their high throughput and economical affordability. For example, microarrays have been applied to studies of cell cycle, regulatory circuitry, cancer cell lines, tumor tissues, and drug discoveries. One obstacle facing the continued success of applying microarray technologies, however, is the random variaton present on microarrays: within signal spots, between spots and among chips. In addition, signals extracted by available software packages seem to vary significantly. Despite a variety of software packages, it appears that there are two major approaches to signal extraction. One approach is to focus on the identification of signal regions and hence estimation of signal levels above background levels. The other approach is to use the distribution of intensity values as a way of identifying relevant signals. Building upon both approaches, the objective of our work is to develop a method that is statistically rigorous and also efficient and robust. Statistical issues to be considered here include: (1) how to refine grid alignment so that the overall variation is minimized, (2) how to estimate the signal levels relative to the local background levels as well as the variance of this estimate, and (3) how to integrate red and green channel signals so that the ratio of interest is stable, simultaneously relaxing distributional assumptions.

NCBI GEO: archive for functional genomics data sets--10 years on.

PubMed

Barrett, Tanya; Troup, Dennis B; Wilhite, Stephen E; Ledoux, Pierre; Evangelista, Carlos; Kim, Irene F; Tomashevsky, Maxim; Marshall, Kimberly A; Phillippy, Katherine H; Sherman, Patti M; Muertter, Rolf N; Holko, Michelle; Ayanbule, Oluwabukunmi; Yefanov, Andrey; Soboleva, Alexandra

2011-01-01

A decade ago, the Gene Expression Omnibus (GEO) database was established at the National Center for Biotechnology Information (NCBI). The original objective of GEO was to serve as a public repository for high-throughput gene expression data generated mostly by microarray technology. However, the research community quickly applied microarrays to non-gene-expression studies, including examination of genome copy number variation and genome-wide profiling of DNA-binding proteins. Because the GEO database was designed with a flexible structure, it was possible to quickly adapt the repository to store these data types. More recently, as the microarray community switches to next-generation sequencing technologies, GEO has again adapted to host these data sets. Today, GEO stores over 20,000 microarray- and sequence-based functional genomics studies, and continues to handle the majority of direct high-throughput data submissions from the research community. Multiple mechanisms are provided to help users effectively search, browse, download and visualize the data at the level of individual genes or entire studies. This paper describes recent database enhancements, including new search and data representation tools, as well as a brief review of how the community uses GEO data. GEO is freely accessible at http://www.ncbi.nlm.nih.gov/geo/.

Development and validation of a microarray for the investigation of the CAZymes encoded by the human gut microbiome.

PubMed

El Kaoutari, Abdessamad; Armougom, Fabrice; Leroy, Quentin; Vialettes, Bernard; Million, Matthieu; Raoult, Didier; Henrissat, Bernard

2013-01-01

Distal gut bacteria play a pivotal role in the digestion of dietary polysaccharides by producing a large number of carbohydrate-active enzymes (CAZymes) that the host otherwise does not produce. We report here the design of a custom microarray that we used to spot non-redundant DNA probes for more than 6,500 genes encoding glycoside hydrolases and lyases selected from 174 reference genomes from distal gut bacteria. The custom microarray was tested and validated by the hybridization of bacterial DNA extracted from the stool samples of lean, obese and anorexic individuals. Our results suggest that a microarray-based study can detect genes from low-abundance bacteria better than metagenomic-based studies. A striking example was the finding that a gene encoding a GH6-family cellulase was present in all subjects examined, whereas metagenomic studies have consistently failed to detect this gene in both human and animal gut microbiomes. In addition, an examination of eight stool samples allowed the identification of a corresponding CAZome core containing 46 families of glycoside hydrolases and polysaccharide lyases, which suggests the functional stability of the gut microbiota despite large taxonomical variations between individuals.

Microarray gene expression profiling analysis combined with bioinformatics in multiple sclerosis.

PubMed

Liu, Mingyuan; Hou, Xiaojun; Zhang, Ping; Hao, Yong; Yang, Yiting; Wu, Xiongfeng; Zhu, Desheng; Guan, Yangtai

2013-05-01

Multiple sclerosis (MS) is the most prevalent demyelinating disease and the principal cause of neurological disability in young adults. Recent microarray gene expression profiling studies have identified several genetic variants contributing to the complex pathogenesis of MS, however, expressional and functional studies are still required to further understand its molecular mechanism. The present study aimed to analyze the molecular mechanism of MS using microarray analysis combined with bioinformatics techniques. We downloaded the gene expression profile of MS from Gene Expression Omnibus (GEO) and analysed the microarray data using the differentially coexpressed genes (DCGs) and links package in R and Database for Annotation, Visualization and Integrated Discovery. The regulatory impact factor (RIF) algorithm was used to measure the impact factor of transcription factor. A total of 1,297 DCGs between MS patients and healthy controls were identified. Functional annotation indicated that these DCGs were associated with immune and neurological functions. Furthermore, the RIF result suggested that IKZF1, BACH1, CEBPB, EGR1, FOS may play central regulatory roles in controlling gene expression in the pathogenesis of MS. Our findings confirm the presence of multiple molecular alterations in MS and indicate the possibility for identifying prognostic factors associated with MS pathogenesis.

Computational synchronization of microarray data with application to Plasmodium falciparum.

PubMed

Zhao, Wei; Dauwels, Justin; Niles, Jacquin C; Cao, Jianshu

2012-06-21

Microarrays are widely used to investigate the blood stage of Plasmodium falciparum infection. Starting with synchronized cells, gene expression levels are continually measured over the 48-hour intra-erythrocytic cycle (IDC). However, the cell population gradually loses synchrony during the experiment. As a result, the microarray measurements are blurred. In this paper, we propose a generalized deconvolution approach to reconstruct the intrinsic expression pattern, and apply it to P. falciparum IDC microarray data. We develop a statistical model for the decay of synchrony among cells, and reconstruct the expression pattern through statistical inference. The proposed method can handle microarray measurements with noise and missing data. The original gene expression patterns become more apparent in the reconstructed profiles, making it easier to analyze and interpret the data. We hypothesize that reconstructed gene expression patterns represent better temporally resolved expression profiles that can be probabilistically modeled to match changes in expression level to IDC transitions. In particular, we identify transcriptionally regulated protein kinases putatively involved in regulating the P. falciparum IDC. By analyzing publicly available microarray data sets for the P. falciparum IDC, protein kinases are ranked in terms of their likelihood to be involved in regulating transitions between the ring, trophozoite and schizont developmental stages of the P. falciparum IDC. In our theoretical framework, a few protein kinases have high probability rankings, and could potentially be involved in regulating these developmental transitions. This study proposes a new methodology for extracting intrinsic expression patterns from microarray data. By applying this method to P. falciparum microarray data, several protein kinases are predicted to play a significant role in the P. falciparum IDC. Earlier experiments have indeed confirmed that several of these kinases are involved in this process. Overall, these results indicate that further functional analysis of these additional putative protein kinases may reveal new insights into how the P. falciparum IDC is regulated.

Detection of cryptic pathogenic copy number variations and constitutional loss of heterozygosity using high resolution SNP microarray analysis in 117 patients referred for cytogenetic analysis and impact on clinical practice.

PubMed

Bruno, D L; Ganesamoorthy, D; Schoumans, J; Bankier, A; Coman, D; Delatycki, M; Gardner, R J M; Hunter, M; James, P A; Kannu, P; McGillivray, G; Pachter, N; Peters, H; Rieubland, C; Savarirayan, R; Scheffer, I E; Sheffield, L; Tan, T; White, S M; Yeung, A; Bowman, Z; Ngo, C; Choy, K W; Cacheux, V; Wong, L; Amor, D J; Slater, H R

2009-02-01

Microarray genome analysis is realising its promise for improving detection of genetic abnormalities in individuals with mental retardation and congenital abnormality. Copy number variations (CNVs) are now readily detectable using a variety of platforms and a major challenge is the distinction of pathogenic from ubiquitous, benign polymorphic CNVs. The aim of this study was to investigate replacement of time consuming, locus specific testing for specific microdeletion and microduplication syndromes with microarray analysis, which theoretically should detect all known syndromes with CNV aetiologies as well as new ones. Genome wide copy number analysis was performed on 117 patients using Affymetrix 250K microarrays. 434 CNVs (195 losses and 239 gains) were found, including 18 pathogenic CNVs and 9 identified as "potentially pathogenic". Almost all pathogenic CNVs were larger than 500 kb, significantly larger than the median size of all CNVs detected. Segmental regions of loss of heterozygosity larger than 5 Mb were found in 5 patients. Genome microarray analysis has improved diagnostic success in this group of patients. Several examples of recently discovered "new syndromes" were found suggesting they are more common than previously suspected and collectively are likely to be a major cause of mental retardation. The findings have several implications for clinical practice. The study revealed the potential to make genetic diagnoses that were not evident in the clinical presentation, with implications for pretest counselling and the consent process. The importance of contributing novel CNVs to high quality databases for genotype-phenotype analysis and review of guidelines for selection of individuals for microarray analysis is emphasised.

ASSESSMENT OF THE SWINE PROTEIN-ANNOTATED OLIGONUCLEOTIDE MICROARRAY AND UTILITY OF THE ARRAYS FOR EQTL AND TRANSCRIPTIONAL PROFILING STUDIES

USDA-ARS?s Scientific Manuscript database

We have evaluated the new Swine Protein-Annotated Oligonucleotide Microarray (http://www.pigoligoarray.org) by analyzing transcriptional profiles for longissimus dorsi muscle (LD), Bronchial lymph node (BLN) and Lung. Four LD samples were used to assess the stringency of hybridization conditions com...

The Microarray Revolution: Perspectives from Educators

ERIC Educational Resources Information Center

Brewster, Jay L.; Beason, K. Beth; Eckdahl, Todd T.; Evans, Irene M.

2004-01-01

In recent years, microarray analysis has become a key experimental tool, enabling the analysis of genome-wide patterns of gene expression. This review approaches the microarray revolution with a focus upon four topics: 1) the early development of this technology and its application to cancer diagnostics; 2) a primer of microarray research,…

High-throughput DNA microarray detection of pathogenic bacteria in shallow well groundwater in the Kathmandu Valley, Nepal.

PubMed

Inoue, Daisuke; Hinoura, Takuji; Suzuki, Noriko; Pang, Junqin; Malla, Rabin; Shrestha, Sadhana; Chapagain, Saroj Kumar; Matsuzawa, Hiroaki; Nakamura, Takashi; Tanaka, Yasuhiro; Ike, Michihiko; Nishida, Kei; Sei, Kazunari

2015-01-01

Because of heavy dependence on groundwater for drinking water and other domestic use, microbial contamination of groundwater is a serious problem in the Kathmandu Valley, Nepal. This study investigated comprehensively the occurrence of pathogenic bacteria in shallow well groundwater in the Kathmandu Valley by applying DNA microarray analysis targeting 941 pathogenic bacterial species/groups. Water quality measurements found significant coliform (fecal) contamination in 10 of the 11 investigated groundwater samples and significant nitrogen contamination in some samples. The results of DNA microarray analysis revealed the presence of 1-37 pathogen species/groups, including 1-27 biosafety level 2 ones, in 9 of the 11 groundwater samples. While the detected pathogens included several feces- and animal-related ones, those belonging to Legionella and Arthrobacter, which were considered not to be directly associated with feces, were detected prevalently. This study could provide a rough picture of overall pathogenic bacterial contamination in the Kathmandu Valley, and demonstrated the usefulness of DNA microarray analysis as a comprehensive screening tool of a wide variety of pathogenic bacteria.

Microarray Analysis of Long Noncoding RNAs in Female Diabetic Peripheral Neuropathy Patients.

PubMed

Luo, Lin; Ji, Lin-Dan; Cai, Jiang-Jia; Feng, Mei; Zhou, Mi; Hu, Su-Pei; Xu, Jin; Zhou, Wen-Hua

2018-01-01

Diabetic peripheral neuropathy (DPN) is the most common complication of diabetes mellitus (DM). Because of its controversial pathogenesis, DPN is still not diagnosed or managed properly in most patients. In this study, human lncRNA microarrays were used to identify the differentially expressed lncRNAs in DM and DPN patients, and some of the discovered lncRNAs were further validated in additional 78 samples by quantitative realtime PCR (qRT-PCR). The microarray analysis identified 446 and 1327 differentially expressed lncRNAs in DM and DPN, respectively. The KEGG pathway analysis further revealed that the differentially expressed lncRNA-coexpressed mRNAs between DPN and DM groups were significantly enriched in the MAPK signaling pathway. The lncRNA/mRNA coexpression network indicated that BDNF and TRAF2 correlated with 6 lncRNAs. The qRT-PCR confirmed the initial microarray results. These findings demonstrated that the interplay between lncRNAs and mRNA may be involved in the pathogenesis of DPN, especially the neurotrophin-MAPK signaling pathway, thus providing relevant information for future studies. © 2018 The Author(s). Published by S. Karger AG, Basel.

Missing value imputation for microarray data: a comprehensive comparison study and a web tool

PubMed Central

2013-01-01

Background Microarray data are usually peppered with missing values due to various reasons. However, most of the downstream analyses for microarray data require complete datasets. Therefore, accurate algorithms for missing value estimation are needed for improving the performance of microarray data analyses. Although many algorithms have been developed, there are many debates on the selection of the optimal algorithm. The studies about the performance comparison of different algorithms are still incomprehensive, especially in the number of benchmark datasets used, the number of algorithms compared, the rounds of simulation conducted, and the performance measures used. Results In this paper, we performed a comprehensive comparison by using (I) thirteen datasets, (II) nine algorithms, (III) 110 independent runs of simulation, and (IV) three types of measures to evaluate the performance of each imputation algorithm fairly. First, the effects of different types of microarray datasets on the performance of each imputation algorithm were evaluated. Second, we discussed whether the datasets from different species have different impact on the performance of different algorithms. To assess the performance of each algorithm fairly, all evaluations were performed using three types of measures. Our results indicate that the performance of an imputation algorithm mainly depends on the type of a dataset but not on the species where the samples come from. In addition to the statistical measure, two other measures with biological meanings are useful to reflect the impact of missing value imputation on the downstream data analyses. Our study suggests that local-least-squares-based methods are good choices to handle missing values for most of the microarray datasets. Conclusions In this work, we carried out a comprehensive comparison of the algorithms for microarray missing value imputation. Based on such a comprehensive comparison, researchers could choose the optimal algorithm for their datasets easily. Moreover, new imputation algorithms could be compared with the existing algorithms using this comparison strategy as a standard protocol. In addition, to assist researchers in dealing with missing values easily, we built a web-based and easy-to-use imputation tool, MissVIA (http://cosbi.ee.ncku.edu.tw/MissVIA), which supports many imputation algorithms. Once users upload a real microarray dataset and choose the imputation algorithms, MissVIA will determine the optimal algorithm for the users' data through a series of simulations, and then the imputed results can be downloaded for the downstream data analyses. PMID:24565220

A Model-Based Joint Identification of Differentially Expressed Genes and Phenotype-Associated Genes

PubMed Central

Seo, Minseok; Shin, Su-kyung; Kwon, Eun-Young; Kim, Sung-Eun; Bae, Yun-Jung; Lee, Seungyeoun; Sung, Mi-Kyung; Choi, Myung-Sook; Park, Taesung

2016-01-01

Over the last decade, many analytical methods and tools have been developed for microarray data. The detection of differentially expressed genes (DEGs) among different treatment groups is often a primary purpose of microarray data analysis. In addition, association studies investigating the relationship between genes and a phenotype of interest such as survival time are also popular in microarray data analysis. Phenotype association analysis provides a list of phenotype-associated genes (PAGs). However, it is sometimes necessary to identify genes that are both DEGs and PAGs. We consider the joint identification of DEGs and PAGs in microarray data analyses. The first approach we used was a naïve approach that detects DEGs and PAGs separately and then identifies the genes in an intersection of the list of PAGs and DEGs. The second approach we considered was a hierarchical approach that detects DEGs first and then chooses PAGs from among the DEGs or vice versa. In this study, we propose a new model-based approach for the joint identification of DEGs and PAGs. Unlike the previous two-step approaches, the proposed method identifies genes simultaneously that are DEGs and PAGs. This method uses standard regression models but adopts different null hypothesis from ordinary regression models, which allows us to perform joint identification in one-step. The proposed model-based methods were evaluated using experimental data and simulation studies. The proposed methods were used to analyze a microarray experiment in which the main interest lies in detecting genes that are both DEGs and PAGs, where DEGs are identified between two diet groups and PAGs are associated with four phenotypes reflecting the expression of leptin, adiponectin, insulin-like growth factor 1, and insulin. Model-based approaches provided a larger number of genes, which are both DEGs and PAGs, than other methods. Simulation studies showed that they have more power than other methods. Through analysis of data from experimental microarrays and simulation studies, the proposed model-based approach was shown to provide a more powerful result than the naïve approach and the hierarchical approach. Since our approach is model-based, it is very flexible and can easily handle different types of covariates. PMID:26964035

Gene expression profile of blood cells for the prediction of delayed cerebral ischemia after intracranial aneurysm rupture: a pilot study in humans.

PubMed

Baumann, Antoine; Devaux, Yvan; Audibert, Gérard; Zhang, Lu; Bracard, Serge; Colnat-Coulbois, Sophie; Klein, Olivier; Zannad, Faiez; Charpentier, Claire; Longrois, Dan; Mertes, Paul-Michel

2013-01-01

Delayed cerebral ischemia (DCI) is a potentially devastating complication after intracranial aneurysm rupture and its mechanisms remain poorly elucidated. Early identification of the patients prone to developing DCI after rupture may represent a major breakthrough in its prevention and treatment. The single gene approach of DCI has demonstrated interest in humans. We hypothesized that whole genome expression profile of blood cells may be useful for better comprehension and prediction of aneurysmal DCI. Over a 35-month period, 218 patients with aneurysm rupture were included in this study. DCI was defined as the occurrence of a new delayed neurological deficit occurring within 2 weeks after aneurysm rupture with evidence of ischemia either on perfusion-diffusion MRI, CT angiography or CT perfusion imaging, or with cerebral angiography. DCI patients were matched against controls based on 4 out of 5 criteria (age, sex, Fisher grade, aneurysm location and smoking status). Genome-wide expression analysis of blood cells obtained at admission was performed by microarrays. Transcriptomic analysis was performed using long oligonucleotide microarrays representing 25,000 genes. Quantitative PCR: 1 µg of total RNA extracted was reverse-transcribed, and the resulting cDNA was diluted 10-fold before performing quantitative PCR. Microarray data were first analyzed by 'Significance Analysis of Microarrays' software which includes the Benjamini correction for multiple testing. In a second step, microarray data fold change was compared using a two-tailed, paired t test. Analysis of receiver-operating characteristic (ROC) curves and the area under the ROC curves were used for prediction analysis. Logistic regression models were used to investigate the additive value of multiple biomarkers. A total of 16 patients demonstrated DCI. Significance Analysis of Microarrays software failed to retrieve significant genes, most probably because of the heterogeneity of the patients included in the microarray experiments and the small size of the DCI population sample. Standard two-tailed paired t test and C-statistic revealed significant associations between gene expression and the occurrence of DCI: in particular, the expression of neuroregulin 1 was 1.6-fold upregulated in patients with DCI (p = 0.01) and predicted DCI with an area under the ROC curve of 0.96. Logistic regression analyses revealed a significant association between neuroregulin 1 and DCI (odds ratio 1.46, 95% confidence interval 1.02-2.09, p = 0.02). This pilot study suggests that blood cells may be a reservoir of prognostic biomarkers of DCI in patients with intracranial aneurysm rupture. Despite an evident lack of power, this study elicited neuroregulin 1, a vasoreactivity-, inflammation- and angiogenesis-related gene, as a possible candidate predictor of DCI. Larger cohort studies are needed but genome-wide microarray-based studies are promising research tools for the understanding of DCI after intracranial aneurysm rupture. © 2013 S. Karger AG, Basel.

A comprehensive sensitivity analysis of microarray breast cancer classification under feature variability

PubMed Central

2009-01-01

Background Large discrepancies in signature composition and outcome concordance have been observed between different microarray breast cancer expression profiling studies. This is often ascribed to differences in array platform as well as biological variability. We conjecture that other reasons for the observed discrepancies are the measurement error associated with each feature and the choice of preprocessing method. Microarray data are known to be subject to technical variation and the confidence intervals around individual point estimates of expression levels can be wide. Furthermore, the estimated expression values also vary depending on the selected preprocessing scheme. In microarray breast cancer classification studies, however, these two forms of feature variability are almost always ignored and hence their exact role is unclear. Results We have performed a comprehensive sensitivity analysis of microarray breast cancer classification under the two types of feature variability mentioned above. We used data from six state of the art preprocessing methods, using a compendium consisting of eight diferent datasets, involving 1131 hybridizations, containing data from both one and two-color array technology. For a wide range of classifiers, we performed a joint study on performance, concordance and stability. In the stability analysis we explicitly tested classifiers for their noise tolerance by using perturbed expression profiles that are based on uncertainty information directly related to the preprocessing methods. Our results indicate that signature composition is strongly influenced by feature variability, even if the array platform and the stratification of patient samples are identical. In addition, we show that there is often a high level of discordance between individual class assignments for signatures constructed on data coming from different preprocessing schemes, even if the actual signature composition is identical. Conclusion Feature variability can have a strong impact on breast cancer signature composition, as well as the classification of individual patient samples. We therefore strongly recommend that feature variability is considered in analyzing data from microarray breast cancer expression profiling experiments. PMID:19941644

Karyotype versus microarray testing for genetic abnormalities after stillbirth.

PubMed

Reddy, Uma M; Page, Grier P; Saade, George R; Silver, Robert M; Thorsten, Vanessa R; Parker, Corette B; Pinar, Halit; Willinger, Marian; Stoll, Barbara J; Heim-Hall, Josefine; Varner, Michael W; Goldenberg, Robert L; Bukowski, Radek; Wapner, Ronald J; Drews-Botsch, Carolyn D; O'Brien, Barbara M; Dudley, Donald J; Levy, Brynn

2012-12-06

Genetic abnormalities have been associated with 6 to 13% of stillbirths, but the true prevalence may be higher. Unlike karyotype analysis, microarray analysis does not require live cells, and it detects small deletions and duplications called copy-number variants. The Stillbirth Collaborative Research Network conducted a population-based study of stillbirth in five geographic catchment areas. Standardized postmortem examinations and karyotype analyses were performed. A single-nucleotide polymorphism array was used to detect copy-number variants of at least 500 kb in placental or fetal tissue. Variants that were not identified in any of three databases of apparently unaffected persons were then classified into three groups: probably benign, clinical significance unknown, or pathogenic. We compared the results of karyotype and microarray analyses of samples obtained after delivery. In our analysis of samples from 532 stillbirths, microarray analysis yielded results more often than did karyotype analysis (87.4% vs. 70.5%, P<0.001) and provided better detection of genetic abnormalities (aneuploidy or pathogenic copy-number variants, 8.3% vs. 5.8%; P=0.007). Microarray analysis also identified more genetic abnormalities among 443 antepartum stillbirths (8.8% vs. 6.5%, P=0.02) and 67 stillbirths with congenital anomalies (29.9% vs. 19.4%, P=0.008). As compared with karyotype analysis, microarray analysis provided a relative increase in the diagnosis of genetic abnormalities of 41.9% in all stillbirths, 34.5% in antepartum stillbirths, and 53.8% in stillbirths with anomalies. Microarray analysis is more likely than karyotype analysis to provide a genetic diagnosis, primarily because of its success with nonviable tissue, and is especially valuable in analyses of stillbirths with congenital anomalies or in cases in which karyotype results cannot be obtained. (Funded by the Eunice Kennedy Shriver National Institute of Child Health and Human Development.).

Towards the integration, annotation and association of historical microarray experiments with RNA-seq.

PubMed

Chavan, Shweta S; Bauer, Michael A; Peterson, Erich A; Heuck, Christoph J; Johann, Donald J

2013-01-01

Transcriptome analysis by microarrays has produced important advances in biomedicine. For instance in multiple myeloma (MM), microarray approaches led to the development of an effective disease subtyping via cluster assignment, and a 70 gene risk score. Both enabled an improved molecular understanding of MM, and have provided prognostic information for the purposes of clinical management. Many researchers are now transitioning to Next Generation Sequencing (NGS) approaches and RNA-seq in particular, due to its discovery-based nature, improved sensitivity, and dynamic range. Additionally, RNA-seq allows for the analysis of gene isoforms, splice variants, and novel gene fusions. Given the voluminous amounts of historical microarray data, there is now a need to associate and integrate microarray and RNA-seq data via advanced bioinformatic approaches. Custom software was developed following a model-view-controller (MVC) approach to integrate Affymetrix probe set-IDs, and gene annotation information from a variety of sources. The tool/approach employs an assortment of strategies to integrate, cross reference, and associate microarray and RNA-seq datasets. Output from a variety of transcriptome reconstruction and quantitation tools (e.g., Cufflinks) can be directly integrated, and/or associated with Affymetrix probe set data, as well as necessary gene identifiers and/or symbols from a diversity of sources. Strategies are employed to maximize the annotation and cross referencing process. Custom gene sets (e.g., MM 70 risk score (GEP-70)) can be specified, and the tool can be directly assimilated into an RNA-seq pipeline. A novel bioinformatic approach to aid in the facilitation of both annotation and association of historic microarray data, in conjunction with richer RNA-seq data, is now assisting with the study of MM cancer biology.

Recent progress in making protein microarray through BioLP

NASA Astrophysics Data System (ADS)

Yang, Rusong; Wei, Lian; Feng, Ying; Li, Xiujian; Zhou, Quan

2017-02-01

Biological laser printing (BioLP) is a promising biomaterial printing technique. It has the advantage of high resolution, high bioactivity, high printing frequency and small transported liquid amount. In this paper, a set of BioLP device is design and made, and protein microarrays are printed by this device. It's found that both laser intensity and fluid layer thickness have an influence on the microarrays acquired. Besides, two kinds of the fluid layer coating methods are compared, and the results show that blade coating method is better than well-coating method in BioLP. A microarray of 0.76pL protein microarray and a "NUDT" patterned microarray are printed to testify the printing ability of BioLP.

Development of a single nucleotide polymorphism DNA microarray for the detection and genotyping of the SARS coronavirus.

PubMed

Guo, Xi; Geng, Peng; Wang, Quan; Cao, Boyang; Liu, Bin

2014-10-01

Severe acute respiratory syndrome (SARS), a disease that spread widely in the world during late 2002 to 2004, severely threatened public health. Although there have been no reported infections since 2004, the extremely pathogenic SARS coronavirus (SARS-CoV), as the causative agent of SARS, has recently been identified in animals, showing the potential for the re-emergence of this disease. Previous studies showed that 27 single nucleotide polymorphism (SNP) mutations among the spike (S) gene of this virus are correlated closely with the SARS pathogenicity and epidemicity. We have developed a SNP DNA microarray in order to detect and genotype these SNPs, and to obtain related information on the pathogenicity and epidemicity of a given strain. The microarray was hybridized with PCR products amplified from cDNAs obtained from different SARS-CoV strains. We were able to detect 24 SNPs and determine the type of a given strain. The hybridization profile showed that 19 samples were detected and genotyped correctly by using our microarray, with 100% accuracy. Our microarray provides a novel method for the detection and epidemiological surveillance of SARS-CoV.

See what you eat--broad GMO screening with microarrays.

PubMed

von Götz, Franz

2010-03-01

Despite the controversy of whether genetically modified organisms (GMOs) are beneficial or harmful for humans, animals, and/or ecosystems, the number of cultivated GMOs is increasing every year. Many countries and federations have implemented safety and surveillance systems for GMOs. Potent testing technologies need to be developed and implemented to monitor the increasing number of GMOs. First, these GMO tests need to be comprehensive, i.e., should detect all, or at least the most important, GMOs on the market. This type of GMO screening requires a high degree of parallel tests or multiplexing. To date, DNA microarrays have the highest number of multiplexing capabilities when nucleic acids are analyzed. This trend article focuses on the evolution of DNA microarrays for GMO testing. Over the last 7 years, combinations of multiplex PCR detection and microarray detection have been developed to qualitatively assess the presence of GMOs. One example is the commercially available DualChip GMO (Eppendorf, Germany; http://www.eppendorf-biochip.com), which is the only GMO screening system successfully validated in a multicenter study. With use of innovative amplification techniques, promising steps have recently been taken to make GMO detection with microarrays quantitative.

Microarray platform for omics analysis

NASA Astrophysics Data System (ADS)

Mecklenburg, Michael; Xie, Bin

2001-09-01

Microarray technology has revolutionized genetic analysis. However, limitations in genome analysis has lead to renewed interest in establishing 'omic' strategies. As we enter the post-genomic era, new microarray technologies are needed to address these new classes of 'omic' targets, such as proteins, as well as lipids and carbohydrates. We have developed a microarray platform that combines self- assembling monolayers with the biotin-streptavidin system to provide a robust, versatile immobilization scheme. A hydrophobic film is patterned on the surface creating an array of tension wells that eliminates evaporation effects thereby reducing the shear stress to which biomolecules are exposed to during immobilization. The streptavidin linker layer makes it possible to adapt and/or develop microarray based assays using virtually any class of biomolecules including: carbohydrates, peptides, antibodies, receptors, as well as them ore traditional DNA based arrays. Our microarray technology is designed to furnish seamless compatibility across the various 'omic' platforms by providing a common blueprint for fabricating and analyzing arrays. The prototype microarray uses a microscope slide footprint patterned with 2 by 96 flat wells. Data on the microarray platform will be presented.

Development of an in vitro skin sensitization test using human cell lines; human Cell Line Activation Test (h-CLAT). II. An inter-laboratory study of the h-CLAT.

PubMed

Sakaguchi, H; Ashikaga, T; Miyazawa, M; Yoshida, Y; Ito, Y; Yoneyama, K; Hirota, M; Itagaki, H; Toyoda, H; Suzuki, H

2006-08-01

Recent regulatory changes have placed a major emphasis on in vitro safety testing and alternative models. In regard to skin sensitization tests, dendritic cells (DCs) derived from human peripheral blood have been considered in the development of new in vitro alternatives. Human cell lines have been also reported recently. In our previous study, we suggested that measuring CD86 and/or CD54 expression on THP-1 cells (human monocytic leukemia cell line) could be used as an in vitro skin sensitization method. An inter-laboratory study among two laboratories was undertaken in Japan in order to further develop an in vitro skin sensitization model. In the present study, we used two human cell lines: THP-1 and U-937 (human histiocytic lymphoma cell line). First we optimized our test protocol (refer to the related paper entitled "optimization of the h-CLAT protocol" within this journal) and then we did an inter-laboratory validation with nine chemicals using the optimized protocol. We measured the expression of CD86 and CD54 on the above cells using flow cytometry after a 24h and 48h exposure to six known allergens (e.g., DNCB, pPD, NiSO(4)) and three non-allergens (e.g., SLS, tween 80). For the sample test concentration, four doses (0.1x, 0.5x, 1x, and 2x of the 50% inhibitory concentration (IC(50))) were evaluated. IC(50) was calculated using MTT assay. We found that allergens/non-allergens were better predicted using THP-1 cells compared to U-937 cells following a 24 h and a 48 h exposure. We also found that the 24h treatment time tended to have a better accuracy than the 48 h treatment time for THP-1 cells. Expression of CD86 and CD54 were good predictive markers for THP-1 cells, but for U-937 cells, expression of CD86 was a better predictor than CD54, at the 24h and the 48 h treatment time. The accuracy also improved when both markers (CD86 and CD54) were used as compared with a single marker for THP-1 cells. Both laboratories gave a good prediction of allergen/non-allergen, especially using THP-1 cells. These results suggest that our method, human Cell Line Activation Test (h-CLAT), using human cell lines THP-1 and U-937, but especially THP-1 cells at 24h treatment, may be a useful in vitro skin sensitization model to predict various contact allergens.

On-Chip Synthesis of Protein Microarrays from DNA Microarrays Via Coupled In Vitro Transcription and Translation for Surface Plasmon Resonance Imaging Biosensor Applications

PubMed Central

Seefeld, Ting H.; Halpern, Aaron R.; Corn, Robert M.

2012-01-01

Protein microarrays are fabricated from double-stranded DNA (dsDNA) microarrays by a one-step, multiplexed enzymatic synthesis in an on-chip microfluidic format and then employed for antibody biosensing measurements with surface plasmon resonance imaging (SPRI). A microarray of dsDNA elements (denoted as generator elements) that encode either a His-tagged green fluorescent protein (GFP) or a His-tagged luciferase protein is utilized to create multiple copies of messenger RNA (mRNA) in a surface RNA polymerase reaction; the mRNA transcripts are then translated into proteins by cell-free protein synthesis in a microfluidic format. The His-tagged proteins diffuse to adjacent Cu(II)-NTA microarray elements (denoted as detector elements) and are specifically adsorbed. The net result is the on-chip, cell-free synthesis of a protein microarray that can be used immediately for SPRI protein biosensing. The dual element format greatly reduces any interference from the nonspecific adsorption of enzyme or proteins. SPRI measurements for the detection of the antibodies anti-GFP and anti-luciferase were used to verify the formation of the protein microarray. This convenient on-chip protein microarray fabrication method can be implemented for multiplexed SPRI biosensing measurements in both clinical and research applications. PMID:22793370

Development and application of a fluorescence protein microarray for detecting serum alpha-fetoprotein in patients with hepatocellular carcinoma.

PubMed

Zhang, Aiying; Yin, Chengzeng; Wang, Zhenshun; Zhang, Yonghong; Zhao, Yuanshun; Li, Ang; Sun, Huanqin; Lin, Dongdong; Li, Ning

2016-12-01

Objective To develop a simple, effective, time-saving and low-cost fluorescence protein microarray method for detecting serum alpha-fetoprotein (AFP) in patients with hepatocellular carcinoma (HCC). Method Non-contact piezoelectric print techniques were applied to fluorescence protein microarray to reduce the cost of prey antibody. Serum samples from patients with HCC and healthy control subjects were collected and evaluated for the presence of AFP using a novel fluorescence protein microarray. To validate the fluorescence protein microarray, serum samples were tested for AFP using an enzyme-linked immunosorbent assay (ELISA). Results A total of 110 serum samples from patients with HCC ( n = 65) and healthy control subjects ( n = 45) were analysed. When the AFP cut-off value was set at 20 ng/ml, the fluorescence protein microarray had a sensitivity of 91.67% and a specificity of 93.24% for detecting serum AFP. Serum AFP quantified via fluorescence protein microarray had a similar diagnostic performance compared with ELISA in distinguishing patients with HCC from healthy control subjects (area under receiver operating characteristic curve: 0.906 for fluorescence protein microarray; 0.880 for ELISA). Conclusion A fluorescence protein microarray method was developed for detecting serum AFP in patients with HCC.

Development and application of a fluorescence protein microarray for detecting serum alpha-fetoprotein in patients with hepatocellular carcinoma

PubMed Central

Zhang, Aiying; Yin, Chengzeng; Wang, Zhenshun; Zhang, Yonghong; Zhao, Yuanshun; Li, Ang; Sun, Huanqin; Lin, Dongdong

2016-01-01

Objective To develop a simple, effective, time-saving and low-cost fluorescence protein microarray method for detecting serum alpha-fetoprotein (AFP) in patients with hepatocellular carcinoma (HCC). Method Non-contact piezoelectric print techniques were applied to fluorescence protein microarray to reduce the cost of prey antibody. Serum samples from patients with HCC and healthy control subjects were collected and evaluated for the presence of AFP using a novel fluorescence protein microarray. To validate the fluorescence protein microarray, serum samples were tested for AFP using an enzyme-linked immunosorbent assay (ELISA). Results A total of 110 serum samples from patients with HCC (n = 65) and healthy control subjects (n = 45) were analysed. When the AFP cut-off value was set at 20 ng/ml, the fluorescence protein microarray had a sensitivity of 91.67% and a specificity of 93.24% for detecting serum AFP. Serum AFP quantified via fluorescence protein microarray had a similar diagnostic performance compared with ELISA in distinguishing patients with HCC from healthy control subjects (area under receiver operating characteristic curve: 0.906 for fluorescence protein microarray; 0.880 for ELISA). Conclusion A fluorescence protein microarray method was developed for detecting serum AFP in patients with HCC. PMID:27885040

Mining microarray data at NCBI's Gene Expression Omnibus (GEO)*.

PubMed

Barrett, Tanya; Edgar, Ron

2006-01-01

The Gene Expression Omnibus (GEO) at the National Center for Biotechnology Information (NCBI) has emerged as the leading fully public repository for gene expression data. This chapter describes how to use Web-based interfaces, applications, and graphics to effectively explore, visualize, and interpret the hundreds of microarray studies and millions of gene expression patterns stored in GEO. Data can be examined from both experiment-centric and gene-centric perspectives using user-friendly tools that do not require specialized expertise in microarray analysis or time-consuming download of massive data sets. The GEO database is publicly accessible through the World Wide Web at http://www.ncbi.nlm.nih.gov/geo.

Performance of the Cottonscan Instrument for Measuring the Average Fiber Linear Density (Fineness) of Cotton Lint Samples

USDA-ARS?s Scientific Manuscript database

This paper explores the CottonscanTM instrument, a new technology designed for routine measurement of the average linear density (fineness) of cotton fiber. A major international inter-laboratory trial of the CottonscanTM system is presented. This expands the range of cottons and laboratories fro...

COMPARISON AND EVALUATION OF LABORATORY PERFORMANCE ON A METHOD FOR THE DETERMINATION OF PERCHLORATE IN FERTILIZERS

EPA Science Inventory

This report details the interlaboratory validation of a method for the determination of perchlorate in fertilizers. In this method (EPA/600/R-01/026), a solid sample of fertilizer is first ground. subsequently, the ground material is either leached with deionized water to dissolv...

OVERVIEW OF AN INTERLABORATORY COLLABORATION ON EVALUATING THE EFFECTS OF MODEL HEPATOTOXICANTS ON HEPATIC GENE EXPRESSION

EPA Science Inventory

Evaluating the Effects of Methapyrilene and Clofibrate on Hepatic Gene Expression: A Collaboration Between Laboratories and a Comparison of Platform and Analytical Approaches

Roger G. Ulrich1, John C. Rockett2, G. Gordon Gibson3 and Syril Pettit4

1 Rosetta Inpharmat...

CHARACTERIZATION OF FINE PARTICLE ASSOCIATED ORGANIC COMPOUNDS: INTERLABORATORY COMPARISON AND DEVELOPMENT OF STANDARD REFERENCE MATERIALS

EPA Science Inventory

Organic chemicals adsorbed to fine particulate matter (PM) in the ambient air account for a major component of the mass and include source tracers as well as toxic compounds that may contribute to adverse human health effects. The US EPA has established a PM 2.5 research progr...

An interlaboratory comparison programme on radio frequency electromagnetic field measurements: the second round of the scheme.

PubMed

Nicolopoulou, E P; Ztoupis, I N; Karabetsos, E; Gonos, I F; Stathopulos, I A

2015-04-01

The second round of an interlaboratory comparison scheme on radio frequency electromagnetic field measurements has been conducted in order to evaluate the overall performance of laboratories that perform measurements in the vicinity of mobile phone base stations and broadcast antenna facilities. The participants recorded the electric field strength produced by two high frequency signal generators inside an anechoic chamber in three measurement scenarios with the antennas transmitting each time different signals at the FM, VHF, UHF and GSM frequency bands. In each measurement scenario, the participants also used their measurements in order to calculate the relative exposure ratios. The results were evaluated in each test level calculating performance statistics (z-scores and En numbers). Subsequently, possible sources of errors for each participating laboratory were discussed, and the overall evaluation of their performances was determined by using an aggregated performance statistic. A comparison between the two rounds proves the necessity of the scheme. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Final report of EURAMET 1197: Supplementary bilateral comparison of hydraulic gauge pressure standards up to 50 MPa

NASA Astrophysics Data System (ADS)

Durgut, Yasin; Petrovski, Nenad; Kacarski, Vanco

2012-01-01

Interlaboratory comparisons are important for the laboratories to assess their own measurement capability. It is equally important for the accreditation bodies and assessors during the audit process of a laboratory to judge whether the laboratory is doing well. As per accreditation rules, it is mandatory for the testing and calibration laboratories to participate in such comparisons from time to time. In this report, results of the bilateral interlaboratory comparison in pressure area in hydraulic media up to 50 MPa gauge between UME (Turkey) and BOM (The FYR of Macedonia) are presented. The artefact used for the comparison was a digital pressure calibrator and its drift was taken into account in the calculation. Results show that all En values lie in acceptable limits. Main text. To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database kcdb.bipm.org/. The final report has been peer-reviewed and approved for publication by EURAMET, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Technical aspects and inter-laboratory variability in native peptide profiling: the CE-MS experience.

PubMed

Mischak, Harald; Vlahou, Antonia; Ioannidis, John P A

2013-04-01

Mass spectrometry platforms have attracted a lot of interest in the last 2 decades as profiling tools for native peptides and proteins with clinical potential. However, limitations associated with reproducibility and analytical robustness, especially pronounced with the initial SELDI systems, hindered the application of such platforms in biomarker qualification and clinical implementation. The scope of this article is to give a short overview on data available on performance and on analytical robustness of the different platforms for peptide profiling. Using the CE-MS platform as a paradigm, data on analytical performance are described including reproducibility (short-term and intermediate repeatability), stability, interference, quantification capabilities (limits of detection), and inter-laboratory variability. We discuss these issues by using as an example our experience with the development of a 273-peptide marker for chronic kidney disease. Finally, we discuss pros and cons and means for improvement and emphasize the need to test in terms of comparative clinical performance and impact, different platforms that pass reasonably well analytical validation tests. Copyright © 2012 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

MicroArray Facility: a laboratory information management system with extended support for Nylon based technologies.

PubMed

Honoré, Paul; Granjeaud, Samuel; Tagett, Rebecca; Deraco, Stéphane; Beaudoing, Emmanuel; Rougemont, Jacques; Debono, Stéphane; Hingamp, Pascal

2006-09-20

High throughput gene expression profiling (GEP) is becoming a routine technique in life science laboratories. With experimental designs that repeatedly span thousands of genes and hundreds of samples, relying on a dedicated database infrastructure is no longer an option.GEP technology is a fast moving target, with new approaches constantly broadening the field diversity. This technology heterogeneity, compounded by the informatics complexity of GEP databases, means that software developments have so far focused on mainstream techniques, leaving less typical yet established techniques such as Nylon microarrays at best partially supported. MAF (MicroArray Facility) is the laboratory database system we have developed for managing the design, production and hybridization of spotted microarrays. Although it can support the widely used glass microarrays and oligo-chips, MAF was designed with the specific idiosyncrasies of Nylon based microarrays in mind. Notably single channel radioactive probes, microarray stripping and reuse, vector control hybridizations and spike-in controls are all natively supported by the software suite. MicroArray Facility is MIAME supportive and dynamically provides feedback on missing annotations to help users estimate effective MIAME compliance. Genomic data such as clone identifiers and gene symbols are also directly annotated by MAF software using standard public resources. The MAGE-ML data format is implemented for full data export. Journalized database operations (audit tracking), data anonymization, material traceability and user/project level confidentiality policies are also managed by MAF. MicroArray Facility is a complete data management system for microarray producers and end-users. Particular care has been devoted to adequately model Nylon based microarrays. The MAF system, developed and implemented in both private and academic environments, has proved a robust solution for shared facilities and industry service providers alike.

MicroArray Facility: a laboratory information management system with extended support for Nylon based technologies

PubMed Central

Honoré, Paul; Granjeaud, Samuel; Tagett, Rebecca; Deraco, Stéphane; Beaudoing, Emmanuel; Rougemont, Jacques; Debono, Stéphane; Hingamp, Pascal

2006-01-01

Background High throughput gene expression profiling (GEP) is becoming a routine technique in life science laboratories. With experimental designs that repeatedly span thousands of genes and hundreds of samples, relying on a dedicated database infrastructure is no longer an option. GEP technology is a fast moving target, with new approaches constantly broadening the field diversity. This technology heterogeneity, compounded by the informatics complexity of GEP databases, means that software developments have so far focused on mainstream techniques, leaving less typical yet established techniques such as Nylon microarrays at best partially supported. Results MAF (MicroArray Facility) is the laboratory database system we have developed for managing the design, production and hybridization of spotted microarrays. Although it can support the widely used glass microarrays and oligo-chips, MAF was designed with the specific idiosyncrasies of Nylon based microarrays in mind. Notably single channel radioactive probes, microarray stripping and reuse, vector control hybridizations and spike-in controls are all natively supported by the software suite. MicroArray Facility is MIAME supportive and dynamically provides feedback on missing annotations to help users estimate effective MIAME compliance. Genomic data such as clone identifiers and gene symbols are also directly annotated by MAF software using standard public resources. The MAGE-ML data format is implemented for full data export. Journalized database operations (audit tracking), data anonymization, material traceability and user/project level confidentiality policies are also managed by MAF. Conclusion MicroArray Facility is a complete data management system for microarray producers and end-users. Particular care has been devoted to adequately model Nylon based microarrays. The MAF system, developed and implemented in both private and academic environments, has proved a robust solution for shared facilities and industry service providers alike. PMID:16987406

The NIST Simple Guide for Evaluating and Expressing Measurement Uncertainty

NASA Astrophysics Data System (ADS)

Possolo, Antonio

2016-11-01

NIST has recently published guidance on the evaluation and expression of the uncertainty of NIST measurement results [1, 2], supplementing but not replacing B. N. Taylor and C. E. Kuyatt's (1994) Guidelines for Evaluating and Expressing the Uncertainty of NIST Measurement Results (NIST Technical Note 1297) [3], which tracks closely the Guide to the expression of uncertainty in measurement (GUM) [4], originally published in 1995 by the Joint Committee for Guides in Metrology of the International Bureau of Weights and Measures (BIPM). The scope of this Simple Guide, however, is much broader than the scope of both NIST Technical Note 1297 and the GUM, because it attempts to address several of the uncertainty evaluation challenges that have arisen at NIST since the 1990s, for example to include molecular biology, greenhouse gases and climate science measurements, and forensic science. The Simple Guide also expands the scope of those two other guidance documents by recognizing observation equations (that is, statistical models) as bona fide measurement models. These models are indispensable to reduce data from interlaboratory studies, to combine measurement results for the same measurand obtained by different methods, and to characterize the uncertainty of calibration and analysis functions used in the measurement of force, temperature, or composition of gas mixtures. This presentation reviews the salient aspects of the Simple Guide, illustrates the use of models and methods for uncertainty evaluation not contemplated in the GUM, and also demonstrates the NIST Uncertainty Machine [5] and the NIST Consensus Builder, which are web-based applications accessible worldwide that facilitate evaluations of measurement uncertainty and the characterization of consensus values in interlaboratory studies.

Detection of clonal evolution in hematopoietic malignancies by combining comparative genomic hybridization and single nucleotide polymorphism arrays.

PubMed

Hartmann, Luise; Stephenson, Christine F; Verkamp, Stephanie R; Johnson, Krystal R; Burnworth, Bettina; Hammock, Kelle; Brodersen, Lisa Eidenschink; de Baca, Monica E; Wells, Denise A; Loken, Michael R; Zehentner, Barbara K

2014-12-01

Array comparative genomic hybridization (aCGH) has become a powerful tool for analyzing hematopoietic neoplasms and identifying genome-wide copy number changes in a single assay. aCGH also has superior resolution compared with fluorescence in situ hybridization (FISH) or conventional cytogenetics. Integration of single nucleotide polymorphism (SNP) probes with microarray analysis allows additional identification of acquired uniparental disomy, a copy neutral aberration with known potential to contribute to tumor pathogenesis. However, a limitation of microarray analysis has been the inability to detect clonal heterogeneity in a sample. This study comprised 16 samples (acute myeloid leukemia, myelodysplastic syndrome, chronic lymphocytic leukemia, plasma cell neoplasm) with complex cytogenetic features and evidence of clonal evolution. We used an integrated manual peak reassignment approach combining analysis of aCGH and SNP microarray data for characterization of subclonal abnormalities. We compared array findings with results obtained from conventional cytogenetic and FISH studies. Clonal heterogeneity was detected in 13 of 16 samples by microarray on the basis of log2 values. Use of the manual peak reassignment analysis approach improved resolution of the sample's clonal composition and genetic heterogeneity in 10 of 13 (77%) patients. Moreover, in 3 patients, clonal disease progression was revealed by array analysis that was not evident by cytogenetic or FISH studies. Genetic abnormalities originating from separate clonal subpopulations can be identified and further characterized by combining aCGH and SNP hybridization results from 1 integrated microarray chip by use of the manual peak reassignment technique. Its clinical utility in comparison to conventional cytogenetic or FISH studies is demonstrated. © 2014 American Association for Clinical Chemistry.

A MICROARRAY ANALYSIS OF GENE EXPRESSION IN THE EMBRYONIC FORELIMB OF THE C57BL/6J MOUSE REVEALS SIGNIFICANT ALTERATIONS METABOLIC AND DEVELOPMENTAL REGULATION FOLLOWING ETHANOL EXPOSURE.

EPA Science Inventory

The observation of transcriptional changes following embryonic ethanol exposure may provide significant insights into the biological response to ethanol exposure. In this study, we used microarray analysis to examine the transcriptional response of the developing limb to a dose ...

Microarrays in brain research: the good, the bad and the ugly.

PubMed

Mirnics, K

2001-06-01

Making sense of microarray data is a complex process, in which the interpretation of findings will depend on the overall experimental design and judgement of the investigator performing the analysis. As a result, differences in tissue harvesting, microarray types, sample labelling and data analysis procedures make post hoc sharing of microarray data a great challenge. To ensure rapid and meaningful data exchange, we need to create some order out of the existing chaos. In these ground-breaking microarray standardization and data sharing efforts, NIH agencies should take a leading role

On the classification techniques in data mining for microarray data classification

NASA Astrophysics Data System (ADS)

Aydadenta, Husna; Adiwijaya

2018-03-01

Cancer is one of the deadly diseases, according to data from WHO by 2015 there are 8.8 million more deaths caused by cancer, and this will increase every year if not resolved earlier. Microarray data has become one of the most popular cancer-identification studies in the field of health, since microarray data can be used to look at levels of gene expression in certain cell samples that serve to analyze thousands of genes simultaneously. By using data mining technique, we can classify the sample of microarray data thus it can be identified with cancer or not. In this paper we will discuss some research using some data mining techniques using microarray data, such as Support Vector Machine (SVM), Artificial Neural Network (ANN), Naive Bayes, k-Nearest Neighbor (kNN), and C4.5, and simulation of Random Forest algorithm with technique of reduction dimension using Relief. The result of this paper show performance measure (accuracy) from classification algorithm (SVM, ANN, Naive Bayes, kNN, C4.5, and Random Forets).The results in this paper show the accuracy of Random Forest algorithm higher than other classification algorithms (Support Vector Machine (SVM), Artificial Neural Network (ANN), Naive Bayes, k-Nearest Neighbor (kNN), and C4.5). It is hoped that this paper can provide some information about the speed, accuracy, performance and computational cost generated from each Data Mining Classification Technique based on microarray data.

ArraySolver: an algorithm for colour-coded graphical display and Wilcoxon signed-rank statistics for comparing microarray gene expression data.

PubMed

Khan, Haseeb Ahmad

2004-01-01

The massive surge in the production of microarray data poses a great challenge for proper analysis and interpretation. In recent years numerous computational tools have been developed to extract meaningful interpretation of microarray gene expression data. However, a convenient tool for two-groups comparison of microarray data is still lacking and users have to rely on commercial statistical packages that might be costly and require special skills, in addition to extra time and effort for transferring data from one platform to other. Various statistical methods, including the t-test, analysis of variance, Pearson test and Mann-Whitney U test, have been reported for comparing microarray data, whereas the utilization of the Wilcoxon signed-rank test, which is an appropriate test for two-groups comparison of gene expression data, has largely been neglected in microarray studies. The aim of this investigation was to build an integrated tool, ArraySolver, for colour-coded graphical display and comparison of gene expression data using the Wilcoxon signed-rank test. The results of software validation showed similar outputs with ArraySolver and SPSS for large datasets. Whereas the former program appeared to be more accurate for 25 or fewer pairs (n < or = 25), suggesting its potential application in analysing molecular signatures that usually contain small numbers of genes. The main advantages of ArraySolver are easy data selection, convenient report format, accurate statistics and the familiar Excel platform.

ArraySolver: An Algorithm for Colour-Coded Graphical Display and Wilcoxon Signed-Rank Statistics for Comparing Microarray Gene Expression Data

PubMed Central

2004-01-01

The massive surge in the production of microarray data poses a great challenge for proper analysis and interpretation. In recent years numerous computational tools have been developed to extract meaningful interpretation of microarray gene expression data. However, a convenient tool for two-groups comparison of microarray data is still lacking and users have to rely on commercial statistical packages that might be costly and require special skills, in addition to extra time and effort for transferring data from one platform to other. Various statistical methods, including the t-test, analysis of variance, Pearson test and Mann–Whitney U test, have been reported for comparing microarray data, whereas the utilization of the Wilcoxon signed-rank test, which is an appropriate test for two-groups comparison of gene expression data, has largely been neglected in microarray studies. The aim of this investigation was to build an integrated tool, ArraySolver, for colour-coded graphical display and comparison of gene expression data using the Wilcoxon signed-rank test. The results of software validation showed similar outputs with ArraySolver and SPSS for large datasets. Whereas the former program appeared to be more accurate for 25 or fewer pairs (n ≤ 25), suggesting its potential application in analysing molecular signatures that usually contain small numbers of genes. The main advantages of ArraySolver are easy data selection, convenient report format, accurate statistics and the familiar Excel platform. PMID:18629036

Differential gene expression detection and sample classification using penalized linear regression models.

PubMed

Wu, Baolin

2006-02-15

Differential gene expression detection and sample classification using microarray data have received much research interest recently. Owing to the large number of genes p and small number of samples n (p > n), microarray data analysis poses big challenges for statistical analysis. An obvious problem owing to the 'large p small n' is over-fitting. Just by chance, we are likely to find some non-differentially expressed genes that can classify the samples very well. The idea of shrinkage is to regularize the model parameters to reduce the effects of noise and produce reliable inferences. Shrinkage has been successfully applied in the microarray data analysis. The SAM statistics proposed by Tusher et al. and the 'nearest shrunken centroid' proposed by Tibshirani et al. are ad hoc shrinkage methods. Both methods are simple, intuitive and prove to be useful in empirical studies. Recently Wu proposed the penalized t/F-statistics with shrinkage by formally using the (1) penalized linear regression models for two-class microarray data, showing good performance. In this paper we systematically discussed the use of penalized regression models for analyzing microarray data. We generalize the two-class penalized t/F-statistics proposed by Wu to multi-class microarray data. We formally derive the ad hoc shrunken centroid used by Tibshirani et al. using the (1) penalized regression models. And we show that the penalized linear regression models provide a rigorous and unified statistical framework for sample classification and differential gene expression detection.

An alternative method to amplify RNA without loss of signal conservation for expression analysis with a proteinase DNA microarray in the ArrayTube format.

PubMed

Schüler, Susann; Wenz, Ingrid; Wiederanders, B; Slickers, P; Ehricht, R

2006-06-12

Recent developments in DNA microarray technology led to a variety of open and closed devices and systems including high and low density microarrays for high-throughput screening applications as well as microarrays of lower density for specific diagnostic purposes. Beside predefined microarrays for specific applications manufacturers offer the production of custom-designed microarrays adapted to customers' wishes. Array based assays demand complex procedures including several steps for sample preparation (RNA extraction, amplification and sample labelling), hybridization and detection, thus leading to a high variability between several approaches and resulting in the necessity of extensive standardization and normalization procedures. In the present work a custom designed human proteinase DNA microarray of lower density in ArrayTube format was established. This highly economic open platform only requires standard laboratory equipment and allows the study of the molecular regulation of cell behaviour by proteinases. We established a procedure for sample preparation and hybridization and verified the array based gene expression profile by quantitative real-time PCR (QRT-PCR). Moreover, we compared the results with the well established Affymetrix microarray. By application of standard labelling procedures with e.g. Klenow fragment exo-, single primer amplification (SPA) or In Vitro Transcription (IVT) we noticed a loss of signal conservation for some genes. To overcome this problem we developed a protocol in accordance with the SPA protocol, in which we included target specific primers designed individually for each spotted oligomer. Here we present a complete array based assay in which only the specific transcripts of interest are amplified in parallel and in a linear manner. The array represents a proof of principle which can be adapted to other species as well. As the designed protocol for amplifying mRNA starts from as little as 100 ng total RNA, it presents an alternative method for detecting even low expressed genes by microarray experiments in a highly reproducible and sensitive manner. Preservation of signal integrity is demonstrated out by QRT-PCR measurements. The little amounts of total RNA necessary for the analyses make this method applicable for investigations with limited material as in clinical samples from, for example, organ or tumour biopsies. Those are arguments in favour of the high potential of our assay compared to established procedures for amplification within the field of diagnostic expression profiling. Nevertheless, the screening character of microarray data must be mentioned, and independent methods should verify the results.

Validation of a two-generational reproduction test in Daphnia magna: An interlaboratory exercise.

PubMed

Barata, Carlos; Campos, Bruno; Rivetti, Claudia; LeBlanc, Gerald A; Eytcheson, Stephanie; McKnight, Stephanie; Tobor-Kaplon, Marysia; de Vries Buitenweg, Selinda; Choi, Suhyon; Choi, Jinhee; Sarapultseva, Elena I; Coutellec, Marie-Agnès; Coke, Maïra; Pandard, Pascal; Chaumot, Arnaud; Quéau, Hervé; Delorme, Nicolas; Geffard, Olivier; Martínez-Jerónimo, Fernando; Watanabe, Haruna; Tatarazako, Norihisa; Lopes, Isabel; Pestana, João L T; Soares, Amadeu M V M; Pereira, Cecilia Manuela; De Schamphelaere, Karel

2017-02-01

Effects observed within one generation disregard potential detrimental effects that may appear across generations. Previously we have developed a two generation Daphnia magna reproduction test using the OECD TG 211 protocol with a few amendments, including initiating the second generation with third brood neonates produced from first generation individuals. Here we showed the results of an inter-laboratory calibration exercise among 12 partners that aimed to test the robustness and consistency of a two generation Daphnia magna reproduction test. Pyperonyl butoxide (PBO) was used as a test compound. Following experiments, PBO residues were determined by TQD-LC/MS/MS. Chemical analysis denoted minor deviations of measured PBO concentrations in freshly prepared and old test solutions and between real and nominal concentrations in all labs. Other test conditions (water, food, D. magna clone, type of test vessel) varied across partners as allowed in the OECD test guidelines. Cumulative fecundity and intrinsic population growth rates (r) were used to estimate "No observed effect concentrations "NOEC using the solvent control as the control treatment. EC 10 and EC- 50 values were obtained regression analyses. Eleven of the twelve labs succeeded in meeting the OECD criteria of producing >60 offspring per female in control treatments during 21days in each of the two consecutive generations. Analysis of variance partitioning of cumulative fecundity indicated a relatively good performance of most labs with most of the variance accounted for by PBO (56.4%) and PBO by interlaboratory interactions (20.2%), with multigenerational effects within and across PBO concentrations explaining about 6% of the variance. EC 50 values for reproduction and population growth rates were on average 16.6 and 20.8% lower among second generation individuals, respectively. In summary these results suggest that the proposed assay is reproducible but cumulative toxicity in the second generation cannot reliably be detected with this assay. Copyright © 2016 Elsevier B.V. All rights reserved.

Interlaboratory comparison for the measurement of particle size and zeta potential of silica nanoparticles in an aqueous suspension

NASA Astrophysics Data System (ADS)

Lamberty, Andrée; Franks, Katrin; Braun, Adelina; Kestens, Vikram; Roebben, Gert; Linsinger, Thomas P. J.

2011-12-01

The Institute for Reference Materials and Measurements has organised an interlaboratory comparison (ILC) to allow the participating laboratories to demonstrate their proficiency in particle size and zeta potential measurements on monomodal aqueous suspensions of silica nanoparticles in the 10-100 nm size range. The main goal of this ILC was to identify competent collaborators for the production of certified nanoparticle reference materials. 38 laboratories from four different continents participated in the ILC with different methods for particle sizing and determination of zeta potential. Most of the laboratories submitted particle size results obtained with centrifugal liquid sedimentation (CLS), dynamic light scattering (DLS) or electron microscopy (EM), or zeta potential values obtained via electrophoretic light scattering (ELS). The results of the laboratories were evaluated using method-specific z scores, calculated on the basis of consensus values from the ILC. For CLS (13 results) and EM (13 results), all reported values were within the ±2 | z| interval. For DLS, 25 of the 27 results reported were within the ±2 | z| interval, the two other results were within the ±3 | z| interval. The standard deviations of the corresponding laboratory mean values varied between 3.7 and 6.5%, which demonstrates satisfactory interlaboratory comparability of CLS, DLS and EM particle size values. From the received test reports, a large discrepancy was observed in terms of the laboratory's quality assurance systems, which are equally important for the selection of collaborators in reference material certification projects. Only a minority of the participating laboratories is aware of all the items that are mandatory in test reports compliant to ISO/IEC 17025 (ISO General requirements for the competence of testing and calibration laboratories. International Organisation for Standardization, Geneva, 2005b). The absence of measurement uncertainty values in the reports, for example, hindered the calculation of zeta scores.

An international collaboration to standardize HIV-2 viral load assays: results from the 2009 ACHI(E)V(2E) quality control study.

PubMed

Damond, F; Benard, A; Balotta, Claudia; Böni, Jürg; Cotten, Matthew; Duque, Vitor; Ferns, Bridget; Garson, Jeremy; Gomes, Perpetua; Gonçalves, Fátima; Gottlieb, Geoffrey; Kupfer, Bernd; Ruelle, Jean; Rodes, Berta; Soriano, Vicente; Wainberg, Mark; Taieb, Audrey; Matheron, Sophie; Chene, Genevieve; Brun-Vezinet, Francoise

2011-10-01

Accurate HIV-2 plasma viral load quantification is crucial for adequate HIV-2 patient management and for the proper conduct of clinical trials and international cohort collaborations. This study compared the homogeneity of HIV-2 RNA quantification when using HIV-2 assays from ACHI(E)V(2E) study sites and either in-house PCR calibration standards or common viral load standards supplied to all collaborators. Each of the 12 participating laboratories quantified blinded HIV-2 samples, using its own HIV-2 viral load assay and standard as well as centrally validated and distributed common HIV-2 group A and B standards (http://www.hiv.lanl.gov/content/sequence/HelpDocs/subtypes-more.html). Aliquots of HIV-2 group A and B strains, each at 2 theoretical concentrations (2.7 and 3.7 log(10) copies/ml), were tested. Intralaboratory, interlaboratory, and overall variances of quantification results obtained with both standards were compared using F tests. For HIV-2 group A quantifications, overall and interlaboratory and/or intralaboratory variances were significantly lower when using the common standard than when using in-house standards at the concentration levels of 2.7 log(10) copies/ml and 3.7 log(10) copies/ml, respectively. For HIV-2 group B, a high heterogeneity was observed and the variances did not differ according to the type of standard used. In this international collaboration, the use of a common standard improved the homogeneity of HIV-2 group A RNA quantification only. The diversity of HIV-2 group B, particularly in PCR primer-binding regions, may explain the heterogeneity in quantification of this strain. Development of a validated HIV-2 viral load assay that accurately quantifies distinct circulating strains is needed.

cSRM 2035: a rare-earth oxide glass for the wavelength calibration of near-infrared dispersive and Fourier transform spectrometers

NASA Astrophysics Data System (ADS)

Choquette, Steven J.; Travis, John C.; Duewer, David L.

1998-10-01

The National Institute of Standards and Technology is developing an optical filter standard for calibration of the wavelength axis of near infrared (NIR) transmission spectrometers. A design goal for the initial candidate Standard Reference Material (cSRM) filter was to provide absorbance peaks evenly covering the spectral region between 800 nm to 1600 mm (12,000 cm-1 to 6,500 cm-1). The reproducibility of the peak location, for batch-certified filters, was to be better than 0.02 nm (approximately 0.1 cm-1). Glasses with 1 to 3 mole % Yb2O3, Sm2O3, and Nd2O3, incorporated into a commercial lanthanum oxide glass were evaluated for this proposed optical standard. An initial batch of cSRM 2035 filters was prepared based on studies of glasses made and evaluated in our laboratory. An interlaboratory comparison study was initiated in February 1997 to evaluate the utility of these filters for the chemical, pharmaceutical, instrumentation, and regulatory communities. Information concerning peak-picking algorithms, wavelength coverage, geometry preferences, and other parameters was solicited from the users. Based upon input from the participants of this interlaboratory study, we are making several changes to make SRM 2035 more useful to our customers. Two of these changes are: (1) incorporating Ho2O3 into the glass to introduce an absorbance peak at approximately 2000 nm (approximately 5000 cm-1) and (2) providing users with a standard center of gravity (COG) peak-picking algorithm to locate the absorbance peaks of the SRM filter precisely. Recent results have demonstrated that the COG method provides a 10 fold improvement in the precision of locating peaks compared with traditional peak-picking methods.

Agreement in assessment of infliximab and adalimumab levels in rheumatoid arthritis: interlaboratory and interassay comparison.

PubMed

Valor, Lara; Hernández-Flórez, Diana; de la Torre, Inmaculada; Llinares, Francisca; Rosas, José; Yagüe, Jordi; Garrido, Jesús; Naredo, Esperanza

2015-01-01

Infliximab (IFX) and adalimumab (ADL) drug levels and anti-drug antibodies (ADA) are assessed using a variety of techniques, therefore, results cannot accurately be compared for clinical purposes. The aim of this study was to test two infliximab (IFX) and adalimumab (ADL) ELISA versions, for drug levels and ADA, to see whether they yield similar results. ELISA versions [Promonitor® IFX R1 and R2 (V.1), Promonitor® IFX and Anti-IFX (V.2); Promonitor® ADL R1 and R2 (V.1), Promonitor® ADL and Anti-ADL (V.2) kits (Progenika Biopharma, Spain)] were used to measure drug levels and ADA in IFX (n=24) and ADL (n=24) rheumatoid arthritis-treated patients in three independent laboratories. Quantitative and qualitative agreements were evaluated using intraclass correlation coefficients (ICC), and Cohen's Kappa (κ) respectively. The Bland-Altman plots assessed differences between V.1 and V.2. Interlaboratory agreement (ICC/κ) with V.1 was poor for IFX (0.66/0.62) and ADL (0.69/0.52) drug levels; meanwhile, high agreement was found with V.2 for IFX (0.98/0.95) and ADL (0.094/1.00). Comparison between V.1 and V.2 in each laboratory resulted in systematically higher values in V.2 than in V.1 and poor agreement (ICC/κ ranges) for IFX (0.12-0.7/ 0.19-0.42) and ADL (0.69-0.89 /0.50-0.73). Qualitative measurements result in better agreement, as evidenced in our study. Greater agreement in V.2 compared with V.1 for IFX and ADL levels could be due to a better tune up. Further studies are required to standardise methods to establish therapeutic reference ranges.

International interlaboratory study comparing single organism 16S rRNA gene sequencing data: Beyond consensus sequence comparisons

PubMed Central

Olson, Nathan D.; Lund, Steven P.; Zook, Justin M.; Rojas-Cornejo, Fabiola; Beck, Brian; Foy, Carole; Huggett, Jim; Whale, Alexandra S.; Sui, Zhiwei; Baoutina, Anna; Dobeson, Michael; Partis, Lina; Morrow, Jayne B.

2015-01-01

This study presents the results from an interlaboratory sequencing study for which we developed a novel high-resolution method for comparing data from different sequencing platforms for a multi-copy, paralogous gene. The combination of PCR amplification and 16S ribosomal RNA gene (16S rRNA) sequencing has revolutionized bacteriology by enabling rapid identification, frequently without the need for culture. To assess variability between laboratories in sequencing 16S rRNA, six laboratories sequenced the gene encoding the 16S rRNA from Escherichia coli O157:H7 strain EDL933 and Listeria monocytogenes serovar 4b strain NCTC11994. Participants performed sequencing methods and protocols available in their laboratories: Sanger sequencing, Roche 454 pyrosequencing®, or Ion Torrent PGM®. The sequencing data were evaluated on three levels: (1) identity of biologically conserved position, (2) ratio of 16S rRNA gene copies featuring identified variants, and (3) the collection of variant combinations in a set of 16S rRNA gene copies. The same set of biologically conserved positions was identified for each sequencing method. Analytical methods using Bayesian and maximum likelihood statistics were developed to estimate variant copy ratios, which describe the ratio of nucleotides at each identified biologically variable position, as well as the likely set of variant combinations present in 16S rRNA gene copies. Our results indicate that estimated variant copy ratios at biologically variable positions were only reproducible for high throughput sequencing methods. Furthermore, the likely variant combination set was only reproducible with increased sequencing depth and longer read lengths. We also demonstrate novel methods for evaluating variable positions when comparing multi-copy gene sequence data from multiple laboratories generated using multiple sequencing technologies. PMID:27077030

High-resolution chromosomal microarrays in prenatal diagnosis significantly increase diagnostic power.

PubMed

Oneda, Beatrice; Baldinger, Rosa; Reissmann, Regina; Reshetnikova, Irina; Krejci, Pavel; Masood, Rahim; Ochsenbein-Kölble, Nicole; Bartholdi, Deborah; Steindl, Katharina; Morotti, Denise; Faranda, Marzia; Baumer, Alessandra; Asadollahi, Reza; Joset, Pascal; Niedrist, Dunja; Breymann, Christian; Hebisch, Gundula; Hüsler, Margaret; Mueller, René; Prentl, Elke; Wisser, Josef; Zimmermann, Roland; Rauch, Anita

2014-06-01

The objective of this study was to determine for the first time the reliability and the diagnostic power of high-resolution microarray testing in routine prenatal diagnostics. We applied high-resolution chromosomal microarray testing in 464 cytogenetically normal prenatal samples with any indication for invasive testing. High-resolution testing revealed a diagnostic yield of 6.9% and 1.6% in cases of fetal ultrasound anomalies and cases of advanced maternal age (AMA), respectively, which is similar to previous studies using low-resolution microarrays. In three (0.6%) additional cases with an indication of AMA, an aberration in susceptibility risk loci was detected. Moreover, one case (0.2%) showed an X-linked aberration in a female fetus, a finding relevant for future family planning. We found the rate of cases, in which the parents had to be tested for interpretation of unreported copy number variants (3.7%), and the rate of remaining variants of unknown significance (0.4%) acceptably low. Of note, these findings did not cause termination of pregnancy after expert genetic counseling. The 0.4% rate of confined placental mosaicism was similar to that observed by conventional karyotyping and notably involved a case of placental microdeletion. High-resolution prenatal microarray testing is a reliable technique that increases diagnostic yield by at least 17.3% when compared with conventional karyotyping, without an increase in the frequency of variants of uncertain significance. © 2014 John Wiley & Sons, Ltd.

Plastic Polymers for Efficient DNA Microarray Hybridization: Application to Microbiological Diagnostics▿

PubMed Central

Zhao, Zhengshan; Peytavi, Régis; Diaz-Quijada, Gerardo A.; Picard, Francois J.; Huletsky, Ann; Leblanc, Éric; Frenette, Johanne; Boivin, Guy; Veres, Teodor; Dumoulin, Michel M.; Bergeron, Michel G.

2008-01-01

Fabrication of microarray devices using traditional glass slides is not easily adaptable to integration into microfluidic systems. There is thus a need for the development of polymeric materials showing a high hybridization signal-to-background ratio, enabling sensitive detection of microbial pathogens. We have developed such plastic supports suitable for highly sensitive DNA microarray hybridizations. The proof of concept of this microarray technology was done through the detection of four human respiratory viruses that were amplified and labeled with a fluorescent dye via a sensitive reverse transcriptase PCR (RT-PCR) assay. The performance of the microarray hybridization with plastic supports made of PMMA [poly(methylmethacrylate)]-VSUVT or Zeonor 1060R was compared to that with high-quality glass slide microarrays by using both passive and microfluidic hybridization systems. Specific hybridization signal-to-background ratios comparable to that obtained with high-quality commercial glass slides were achieved with both polymeric substrates. Microarray hybridizations demonstrated an analytical sensitivity equivalent to approximately 100 viral genome copies per RT-PCR, which is at least 100-fold higher than the sensitivities of previously reported DNA hybridizations on plastic supports. Testing of these plastic polymers using a microfluidic microarray hybridization platform also showed results that were comparable to those with glass supports. In conclusion, PMMA-VSUVT and Zeonor 1060R are both suitable for highly sensitive microarray hybridizations. PMID:18784318

Development and application of a microarray meter tool to optimize microarray experiments

PubMed Central

Rouse, Richard JD; Field, Katrine; Lapira, Jennifer; Lee, Allen; Wick, Ivan; Eckhardt, Colleen; Bhasker, C Ramana; Soverchia, Laura; Hardiman, Gary

2008-01-01

Background Successful microarray experimentation requires a complex interplay between the slide chemistry, the printing pins, the nucleic acid probes and targets, and the hybridization milieu. Optimization of these parameters and a careful evaluation of emerging slide chemistries are a prerequisite to any large scale array fabrication effort. We have developed a 'microarray meter' tool which assesses the inherent variations associated with microarray measurement prior to embarking on large scale projects. Findings The microarray meter consists of nucleic acid targets (reference and dynamic range control) and probe components. Different plate designs containing identical probe material were formulated to accommodate different robotic and pin designs. We examined the variability in probe quality and quantity (as judged by the amount of DNA printed and remaining post-hybridization) using three robots equipped with capillary printing pins. Discussion The generation of microarray data with minimal variation requires consistent quality control of the (DNA microarray) manufacturing and experimental processes. Spot reproducibility is a measure primarily of the variations associated with printing. The microarray meter assesses array quality by measuring the DNA content for every feature. It provides a post-hybridization analysis of array quality by scoring probe performance using three metrics, a) a measure of variability in the signal intensities, b) a measure of the signal dynamic range and c) a measure of variability of the spot morphologies. PMID:18710498

A Web-Based Multi-Database System Supporting Distributed Collaborative Management and Sharing of Microarray Experiment Information

PubMed Central

Burgarella, Sarah; Cattaneo, Dario; Masseroli, Marco

2006-01-01

We developed MicroGen, a multi-database Web based system for managing all the information characterizing spotted microarray experiments. It supports information gathering and storing according to the Minimum Information About Microarray Experiments (MIAME) standard. It also allows easy sharing of information and data among all multidisciplinary actors involved in spotted microarray experiments. PMID:17238488

Polysaccharide Microarray Technology for the Detection of Burkholderia Pseudomallei and Burkholderia Mallei Antibodies

DTIC Science & Technology

2006-04-27

polysaccharide microarray platform was prepared by immobilizing Burkholderia pseudomallei and Burkholderia mallei polysaccharides . This... polysaccharide array was tested with success for detecting B. pseudomallei and B. mallei serum (human and animal) antibodies. The advantages of this microarray... Polysaccharide microarrays; Burkholderia pseudomallei; Burkholderia mallei; Glanders; Melioidosis1. Introduction There has been a great deal of emphasis on the

Microarray-integrated optoelectrofluidic immunoassay system

PubMed Central

Han, Dongsik

2016-01-01

A microarray-based analytical platform has been utilized as a powerful tool in biological assay fields. However, an analyte depletion problem due to the slow mass transport based on molecular diffusion causes low reaction efficiency, resulting in a limitation for practical applications. This paper presents a novel method to improve the efficiency of microarray-based immunoassay via an optically induced electrokinetic phenomenon by integrating an optoelectrofluidic device with a conventional glass slide-based microarray format. A sample droplet was loaded between the microarray slide and the optoelectrofluidic device on which a photoconductive layer was deposited. Under the application of an AC voltage, optically induced AC electroosmotic flows caused by a microarray-patterned light actively enhanced the mass transport of target molecules at the multiple assay spots of the microarray simultaneously, which reduced tedious reaction time from more than 30 min to 10 min. Based on this enhancing effect, a heterogeneous immunoassay with a tiny volume of sample (5 μl) was successfully performed in the microarray-integrated optoelectrofluidic system using immunoglobulin G (IgG) and anti-IgG, resulting in improved efficiency compared to the static environment. Furthermore, the application of multiplex assays was also demonstrated by multiple protein detection. PMID:27190571

Microarray-integrated optoelectrofluidic immunoassay system.

PubMed

Han, Dongsik; Park, Je-Kyun

2016-05-01

A microarray-based analytical platform has been utilized as a powerful tool in biological assay fields. However, an analyte depletion problem due to the slow mass transport based on molecular diffusion causes low reaction efficiency, resulting in a limitation for practical applications. This paper presents a novel method to improve the efficiency of microarray-based immunoassay via an optically induced electrokinetic phenomenon by integrating an optoelectrofluidic device with a conventional glass slide-based microarray format. A sample droplet was loaded between the microarray slide and the optoelectrofluidic device on which a photoconductive layer was deposited. Under the application of an AC voltage, optically induced AC electroosmotic flows caused by a microarray-patterned light actively enhanced the mass transport of target molecules at the multiple assay spots of the microarray simultaneously, which reduced tedious reaction time from more than 30 min to 10 min. Based on this enhancing effect, a heterogeneous immunoassay with a tiny volume of sample (5 μl) was successfully performed in the microarray-integrated optoelectrofluidic system using immunoglobulin G (IgG) and anti-IgG, resulting in improved efficiency compared to the static environment. Furthermore, the application of multiplex assays was also demonstrated by multiple protein detection.

Carbohydrate Microarray Technology Applied to High-Throughput Mapping of Plant Cell Wall Glycans Using Comprehensive Microarray Polymer Profiling (CoMPP).

PubMed

Kračun, Stjepan Krešimir; Fangel, Jonatan Ulrik; Rydahl, Maja Gro; Pedersen, Henriette Lodberg; Vidal-Melgosa, Silvia; Willats, William George Tycho

2017-01-01

Cell walls are an important feature of plant cells and a major component of the plant glycome. They have both structural and physiological functions and are critical for plant growth and development. The diversity and complexity of these structures demand advanced high-throughput techniques to answer questions about their structure, functions and roles in both fundamental and applied scientific fields. Microarray technology provides both the high-throughput and the feasibility aspects required to meet that demand. In this chapter, some of the most recent microarray-based techniques relating to plant cell walls are described together with an overview of related contemporary techniques applied to carbohydrate microarrays and their general potential in glycoscience. A detailed experimental procedure for high-throughput mapping of plant cell wall glycans using the comprehensive microarray polymer profiling (CoMPP) technique is included in the chapter and provides a good example of both the robust and high-throughput nature of microarrays as well as their applicability to plant glycomics.

Mutation analysis of 272 Spanish families affected by autosomal recessive retinitis pigmentosa using a genotyping microarray.

PubMed

Ávila-Fernández, Almudena; Cantalapiedra, Diego; Aller, Elena; Vallespín, Elena; Aguirre-Lambán, Jana; Blanco-Kelly, Fiona; Corton, M; Riveiro-Álvarez, Rosa; Allikmets, Rando; Trujillo-Tiebas, María José; Millán, José M; Cremers, Frans P M; Ayuso, Carmen

2010-12-03

Retinitis pigmentosa (RP) is a genetically heterogeneous disorder characterized by progressive loss of vision. The aim of this study was to identify the causative mutations in 272 Spanish families using a genotyping microarray. 272 unrelated Spanish families, 107 with autosomal recessive RP (arRP) and 165 with sporadic RP (sRP), were studied using the APEX genotyping microarray. The families were also classified by clinical criteria: 86 juveniles and 186 typical RP families. Haplotype and sequence analysis were performed to identify the second mutated allele. At least one-gene variant was found in 14% and 16% of the juvenile and typical RP groups respectively. Further study identified four new mutations, providing both causative changes in 11% of the families. Retinol Dehydrogenase 12 (RDH12) was the most frequently mutated gene in the juvenile RP group, and Usher Syndrome 2A (USH2A) and Ceramide Kinase-Like (CERKL) were the most frequently mutated genes in the typical RP group. The only variant found in CERKL was p.Arg257Stop, the most frequent mutation. The genotyping microarray combined with segregation and sequence analysis allowed us to identify the causative mutations in 11% of the families. Due to the low number of characterized families, this approach should be used in tandem with other techniques.

NCBI GEO: archive for functional genomics data sets—10 years on

PubMed Central

Barrett, Tanya; Troup, Dennis B.; Wilhite, Stephen E.; Ledoux, Pierre; Evangelista, Carlos; Kim, Irene F.; Tomashevsky, Maxim; Marshall, Kimberly A.; Phillippy, Katherine H.; Sherman, Patti M.; Muertter, Rolf N.; Holko, Michelle; Ayanbule, Oluwabukunmi; Yefanov, Andrey; Soboleva, Alexandra

2011-01-01

A decade ago, the Gene Expression Omnibus (GEO) database was established at the National Center for Biotechnology Information (NCBI). The original objective of GEO was to serve as a public repository for high-throughput gene expression data generated mostly by microarray technology. However, the research community quickly applied microarrays to non-gene-expression studies, including examination of genome copy number variation and genome-wide profiling of DNA-binding proteins. Because the GEO database was designed with a flexible structure, it was possible to quickly adapt the repository to store these data types. More recently, as the microarray community switches to next-generation sequencing technologies, GEO has again adapted to host these data sets. Today, GEO stores over 20 000 microarray- and sequence-based functional genomics studies, and continues to handle the majority of direct high-throughput data submissions from the research community. Multiple mechanisms are provided to help users effectively search, browse, download and visualize the data at the level of individual genes or entire studies. This paper describes recent database enhancements, including new search and data representation tools, as well as a brief review of how the community uses GEO data. GEO is freely accessible at http://www.ncbi.nlm.nih.gov/geo/. PMID:21097893

Clinical application of chromosomal microarray analysis for the prenatal diagnosis of chromosomal abnormalities and copy number variations in fetuses with congenital heart disease.

PubMed

Xia, Yu; Yang, Yongchao; Huang, Shufang; Wu, Yueheng; Li, Ping; Zhuang, Jian

2018-03-24

This study aimed to determine chromosomal abnormalities and copy number variations (CNVs) in fetuses with congenital heart disease (CHD) by chromosomal microarray analysis (CMA). One hundred and ten cases with CHD detected by prenatal echocardiography were enrolled in the study; 27 cases were simple CHDs, and 83 were complex CHDs. Chromosomal microarray analysis was performed on the Affymetrix CytoScan HD platform. All annotated CNVs were validated by quantitative PCR. Chromosomal microarray analysis identified 6 cases with chromosomal abnormalities, including 2 cases with trisomy 21, 2 cases with trisomy 18, 1 case with trisomy 13, and 1 unusual case of mosaic trisomy 21. Pathogenic CNVs were detected in 15.5% (17/110) of the fetuses with CHDs, including 13 cases with CHD-associated CNVs. We further identified 10 genes as likely novel CHD candidate genes through gene functional enrichment analysis. We also found that pathogenic CMA results impacted the rate of pregnancy termination. This study shows that CMA is particularly effective for identifying chromosomal abnormalities and CNVs in fetuses with CHDs as well as having an effect on obstetrical outcomes. The elucidation of the genetic basis of CHDs will continue to expand our understanding of the etiology of CHDs. © 2018 John Wiley & Sons, Ltd.

Interim report on updated microarray probes for the LLNL Burkholderia pseudomallei SNP array

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gardner, S; Jaing, C

2012-03-27

The overall goal of this project is to forensically characterize 100 unknown Burkholderia isolates in the US-Australia collaboration. We will identify genome-wide single nucleotide polymorphisms (SNPs) from B. pseudomallei and near neighbor species including B. mallei, B. thailandensis and B. oklahomensis. We will design microarray probes to detect these SNP markers and analyze 100 Burkholderia genomic DNAs extracted from environmental, clinical and near neighbor isolates from Australian collaborators on the Burkholderia SNP microarray. We will analyze the microarray genotyping results to characterize the genetic diversity of these new isolates and triage the samples for whole genome sequencing. In this interimmore » report, we described the SNP analysis and the microarray probe design for the Burkholderia SNP microarray.« less

Using in vitro models for expression profiling studies on ethanol and drugs of abuse.

PubMed

Thibault, Christelle; Hassan, Sajida; Miles, Michael

2005-03-01

The use of expression profiling with microarrays offers great potential for studying the mechanisms of action of drugs of abuse. Studies with the intact nervous system seem likely to be most relevant to understanding the mechanisms of drug abuse-related behaviours. However, the use of expression profiling with in vitro culture models offers significant advantages for identifying details of cellular signalling actions and toxicity for drugs of abuse. This study discusses general issues of the use of microarrays and cell culture models for studies on drugs of abuse. Specific results from existing studies are also discussed, providing clear examples of relevance for in vitro studies on ethanol, nicotine, opiates, cannabinoids and hallucinogens such as LSD. In addition to providing details on signalling mechanisms relevant to the neurobiology of drugs of abuse, microarray studies on a variety of cell culture systems have also provided important information on mechanisms of cellular/organ toxicity with drugs of abuse. Efforts to integrate genomic studies on drugs of abuse with both in vivo and in vitro models offer the potential for novel mechanistic rigor and physiological relevance.

Transcriptome Profiling of In-Vivo Produced Bovine Pre-implantation Embryos Using Two-color Microarray Platform.

PubMed

Salehi, Reza; Tsoi, Stephen C M; Colazo, Marcos G; Ambrose, Divakar J; Robert, Claude; Dyck, Michael K

2017-01-30

Early embryonic loss is a large contributor to infertility in cattle. Moreover, bovine becomes an interesting model to study human preimplantation embryo development due to their similar developmental process. Although genetic factors are known to affect early embryonic development, the discovery of such factors has been a serious challenge. Microarray technology allows quantitative measurement and gene expression profiling of transcript levels on a genome-wide basis. One of the main decisions that have to be made when planning a microarray experiment is whether to use a one- or two-color approach. Two-color design increases technical replication, minimizes variability, improves sensitivity and accuracy as well as allows having loop designs, defining the common reference samples. Although microarray is a powerful biological tool, there are potential pitfalls that can attenuate its power. Hence, in this technical paper we demonstrate an optimized protocol for RNA extraction, amplification, labeling, hybridization of the labeled amplified RNA to the array, array scanning and data analysis using the two-color analysis strategy.

Metadata management and semantics in microarray repositories.

PubMed

Kocabaş, F; Can, T; Baykal, N

2011-12-01

The number of microarray and other high-throughput experiments on primary repositories keeps increasing as do the size and complexity of the results in response to biomedical investigations. Initiatives have been started on standardization of content, object model, exchange format and ontology. However, there are backlogs and inability to exchange data between microarray repositories, which indicate that there is a great need for a standard format and data management. We have introduced a metadata framework that includes a metadata card and semantic nets that make experimental results visible, understandable and usable. These are encoded in syntax encoding schemes and represented in RDF (Resource Description Frame-word), can be integrated with other metadata cards and semantic nets, and can be exchanged, shared and queried. We demonstrated the performance and potential benefits through a case study on a selected microarray repository. We concluded that the backlogs can be reduced and that exchange of information and asking of knowledge discovery questions can become possible with the use of this metadata framework.