Interlaboratory Data Research Articles

Evaluation of extraction and storage conditions for quantification and characterization of silver nanoparticles in complex samples by single particle-ICP-MS

The extraction of nanoparticles (NPs) from complex matrices and subsequent storage can potentially alter the NPs physicochemical properties and hinder cross-study comparisons. Most NP extraction methods are designed and tested at high nanoparticle concentrations, although (eco)toxicological and regulatory monitoring programs require methods capable of analyzing NPs at environmentally relevant concentrations (lower ppb range). In this study, we investigated how extraction methods affect the characteristics of PVP coated and citrate-stabilized silver NPs (AgNPs) spiked into soil, sewage sludge, and biological samples at environmentally relevant concentrations using Single Particle Inductively Coupled Plasma Mass Spectrometry spICP-MS). Further we investigated the impact of storage temperature (-80°C to 21°C) and storage duration (1-28 days) on the particle characteristics such as particle size.We found that aqueous AgNPs samples with low ionic strength media retained their original characteristics (like particle size, particle concentration and particle-based Ag mass) when preserved at 4°C for up to 28 days. AgNPs dispersed in high ionic strength media were however better preserved at -80°C. Among the extraction agents, tetrasodium pyrophosphate was more efficient in extracting AgNPs from soil and sewage sludge matrices, while Proteinase K was more suitable for biological samples from organisms (earthworms or fish).Although our study focused only on AgNPs, it provides crucial information to aid interlaboratory comparisons and data interpretation for (eco)toxicological studies.

Application of pulsed-field gel electrophoresis for molecular identification of pathogenic Leptospira species in Iran: a rapid and reliable method.

Leptospirosis is a zoonotic disease caused by pathogenic Leptospira serovars. The genus Leptospira cannot differentiated by conventional techniques. However, identity determination of pathogenic serovar is precious of public health problems and epidemiological studies. Pulsed-field gel electrophoresis facilitates rapid identification of Leptospires to the serovar levels. In this study, we employed PFGE to evaluate 28 Leptospira isolates, with animal, human and environmental origin, obtained from Razi Vaccine and Serum Research Institute of Karaj, Iran. PFGE patterns of 28 Leptospira serovars were generated using the Not I restriction enzyme in comparison with the lambda ladder. Out of 28 serovars evaluated, we identified 22 different pulsed types, designated P1-P22. Out of 22 pulse groups, 3 were found to be a common type, but others were a single Type. Groups consisting of the common type were P3, P9, P14, and P16. The results showed that the discriminatory index of PFGE by Not I enzyme was 0.99, demonstrating heterogeneous differentiation among serovar members. The PFGE methodology used in this study showed excellent interlaboratory report usability, rapid, reliable, enabling standardization and data sharing between laboratories.

A template wizard for the cocreation of machine-readable data-reporting to harmonize the evaluation of (nano)materials.

Making research data findable, accessible, interoperable and reusable (FAIR) is typically hampered by a lack of skills in technical aspects of data management by data generators and a lack of resources. We developed a Template Wizard for researchers to easily create templates suitable for consistently capturing data and metadata from their experiments. The templates are easy to use and enable the compilation of machine-readable metadata to accompany data generation and align them to existing community standards and databases, such as eNanoMapper, streamlining the adoption of the FAIR principles. These templates are citable objects and are available as online tools. The Template Wizard is designed to be user friendly and facilitates using and reusing existing templates for new projects or project extensions. The wizard is accompanied by an online template validator, which allows self-evaluation of the template (to ensure mapping to the data schema and machine readability of the captured data) and transformation by an open-source parser into machine-readable formats, compliant with the FAIR principles. The templates are based on extensive collective experience in nanosafety data collection and include over 60 harmonized data entry templates for physicochemical characterization and hazard assessment (cell viability, genotoxicity, environmental organism dose-response tests, omics), as well as exposure and release studies. The templates are generalizable across fields and have already been extended and adapted for microplastics and advanced materials research. The harmonized templates improve the reliability of interlaboratory comparisons, data reuse and meta-analyses and can facilitate the safety evaluation and regulation process for (nano) materials.

Quartet metabolite reference materials for inter-laboratory proficiency test and data integration of metabolomics profiling

BackgroundVarious laboratory-developed metabolomic methods lead to big challenges in inter-laboratory comparability and effective integration of diverse datasets.ResultsAs part of the Quartet Project, we establish a publicly available suite of four metabolite reference materials derived from B lymphoblastoid cell lines from a family of parents and monozygotic twin daughters. We generate comprehensive LC–MS-based metabolomic data from the Quartet reference materials using targeted and untargeted strategies in different laboratories. The Quartet multi-sample-based signal-to-noise ratio enables objective assessment of the reliability of intra-batch and cross-batch metabolomics profiling in detecting intrinsic biological differences among the four groups of samples. Significant variations in the reliability of the metabolomics profiling are identified across laboratories. Importantly, ratio-based metabolomics profiling, by scaling the absolute values of a study sample relative to those of a common reference sample, enables cross-laboratory quantitative data integration. Thus, we construct the ratio-based high-confidence reference datasets between two reference samples, providing “ground truth” for inter-laboratory accuracy assessment, which enables objective evaluation of quantitative metabolomics profiling using various instruments and protocols.ConclusionsOur study provides the community with rich resources and best practices for inter-laboratory proficiency tests and data integration, ensuring reliability of large-scale and longitudinal metabolomic studies.

ECerto—versatile software for interlaboratory data evaluation and documentation during reference material production

The statistical tool eCerto was developed for the evaluation of measurement data to assign property values and associated uncertainties of reference materials. The analysis is based on collaborative studies of expert laboratories and was implemented using the R software environment. Emphasis was put on comparability of eCerto with SoftCRM, a statistical tool based on the certification strategy of the former Community Bureau of Reference. Additionally, special attention was directed towards easy usability from data collection through processing, archiving, and reporting. While the effects of outlier removal can be flexibly explored, eCerto always retains the original data set and any manipulation such as outlier removal is (graphically and tabularly) documented adequately in the report. As a major reference materials producer, the Bundesanstalt für Materialforschung und -prüfung (BAM) developed and will maintain a tool to meet the needs of modern data processing, documentation requirements, and emerging fields of RM activity. The main features of eCerto are discussed using previously certified reference materials.Graphical

50 Human Hazard Assessment of Nanomaterials: Supporting Risk Decision Making Through Interlaboratory Trial Data

Abstract The prevalent application of engineered nanomaterials (ENMs) has led to an extensive effort to develop tools facilitating their risk assessment and management. This interlaboratory trial (encompassing two laboratories) sought to investigate the genotoxic impact of ENMs using the hypoxanthine phosphoribosyltransferase (HPRT) forward mutation assay and in vitro cytokinesis-blocked micronucleus (CBMN) assay, utilising harmonised protocols. In the CBMN assay, human lymphoblast (TK6) cells were exposed to zinc oxide (ZnO), titanium dioxide (TiO2) and tungsten carbide-cobalt (WC/Co) for 24-hours after which 1000 binucleated cells were scored for micronuclei (n=2). For the HPRT assay, TK6 and mouse fibroblast (V79) cells were exposed to ZnO, TiO2, WC/Co, CuO and Nanocyl-CNTs for 24-hours after which 600 wells were scored for point mutations (n=2). Significant cytotoxicity and micronuclei frequency was reported following ZnO exposure (20µg/ml). Significant micronuclei induction was reported following WC/Co exposures (2-fold over control, 100µg/ml). No significant mutagenicity was detected with ZnO or TiO2; both laboratories observed significant cytotoxicity following exposure to ZnO (20µg/ml; 37% reduction in cell viability). In the second interlaboratory trial CuO induced significant cytotoxicity (36% reduction in viability: 0.5µg/ml), and a 32-fold increase in mutagenicity. The Nanocyl induced a 6-fold increase in mutagenicity. The data generated here has shown good statistical concordance between laboratories and has highlighted a promising route forward in supporting risk decision making for ENMs. The authors would like to acknowledge this research has received funding from the European Union’s Horizon 2020 research and innovation program for the RiskGONE project, grant agreement #814425.

SMART: A data reporting standard for mass spectrometry imaging.

Mass spectrometry imaging (MSI) is an important analytical technique that simultaneously reports the spatial location and abundance of detected ions in biological, chemical, clinical, and pharmaceutical studies. As MSI grows in popularity, it has become evident that data reporting varies among different research groups and between techniques. The lack of consistency in data reporting inherently creates additional challenges in comparing intra- and inter-laboratory MSI data. In this tutorial, we propose a unified data reporting system, SMART, based on the common features shared between techniques. While there are limitations to any reporting system, SMART was decided upon after significant discussion to more easily understand and benchmark MSI data. SMART is not intended to be comprehensive but rather capture essential baseline information for a given MSI study; this could be within a study (e.g., effect of spot size on the measured ion signals) or between two studies (e.g., different MSI platform technologies applied to the same tissue type). This tutorial does not attempt to address the confidence with which annotations are made nor does it deny the importance of other parameters that are not included in the current SMART format. Ultimately, the goal of this tutorial is to discuss the necessity of establishing a uniform reporting system to communicate MSI data in publications and presentations in a simple format to readily interpret the parameters and baseline outcomes of the data.

Experimental factors that impact CaV1.2 channel pharmacology-Effects of recording temperature, charge carrier, and quantification of drug effects on the step and ramp currents elicited by the "step-step-ramp" voltage protocol.

CaV1.2 channels contribute to action potential upstroke in pacemaker cells, plateau potential in working myocytes, and initiate excitation-contraction coupling. Understanding drug action on CaV1.2 channels may inform potential impact on cardiac function. However, literature shows large degrees of variability between CaV1.2 pharmacology generated by different laboratories, casting doubt regarding the utility of these data to predict or interpret clinical outcomes. This study examined experimental factors that may impact CaV1.2 pharmacology. Whole cell recordings were made on CaV1.2 overexpression cells. Current was evoked using a "step-step-ramp" waveform that elicited a step and a ramp current. Experimental factors examined were: 1) near physiological vs. room temperature for recording, 2) drug inhibition of the step vs. the ramp current, and 3) Ca2+ vs. Ba2+ as the charge carrier. Eight drugs were studied. CaV1.2 current exhibited prominent rundown, exquisite temperature sensitivity, and required a high degree of series resistance compensation to optimize voltage control. Temperature-dependent effects were examined for verapamil and methadone. Verapamil's block potency shifted by up to 4X between room to near physiological temperature. Methadone exhibited facilitatory and inhibitory effects at near physiological temperature, and only inhibitory effect at room temperature. Most drugs inhibited the ramp current more potently than the step current-a preference enhanced when Ba2+ was the charge carrier. The slopes of the concentration-inhibition relationships for many drugs were shallow, temperature-dependent, and differed between the step and the ramp current. All experimental factors examined affected CaV1.2 pharmacology. In addition, whole cell CaV1.2 current characteristics-rundown, temperature sensitivity, and impact of series resistance-are also factors that can impact pharmacology. Drug effects on CaV1.2 channels appear more complex than simple pore block mechanism. Normalizing laboratory-specific approaches is key to improve inter-laboratory data reproducibility. Releasing original electrophysiology records is essential to promote transparency and enable the independent evaluation of data quality.

Validation of the Assurance® GDS for Cronobacter Tq II in Infant Formulas, Infant Cereals, Ingredients, and Environmental Samples Collaborative Study: First Action 2021.08.

The Assurance® GDS for Cronobacter Tq II assay is a nucleic acid amplification system for the qualitative detection of Cronobacter. The method uses an upfront concentration of the target organism from the enrichment by immunomagnetic separation (IMS) using the PickPen®device. The Assurance GDS for Cronobacter Tq II method was evaluated for Official Methods of AnalysisSM certification. The matrix was compared to the ISO 22964:2017: Microbiology of the Food Chain-Horizontal Method for the Detection of Cronobacter spp. standard and using an alternative confirmation procedure. The alternative method was evaluated using 10 g test portions in an unpaired study design for powdered infant formula (milk based with iron and DHA) containing probiotics. Eleven technicians from eight laboratories located within the United States and Europe participated in the collaborative study. Statistical analysis was conducted according to the probability of detection (POD) statistical model as presented in the AOAC validation guidelines. The difference in laboratory POD (dLPODC) values with 95% confidence intervals across collaborators was calculated for each level between the candidate and reference method results and between the candidate presumptive and confirmed results. Results obtained for the low inoculum level test portions produced a dLPOD value with a 95% confidence interval of 0.03 (-0.18, 0.15). The dLPOD results indicate equivalence between the candidate method and reference method for the matrix evaluated. The method also demonstrated acceptable inter-laboratory reproducibility as determined in the collaborative evaluation. There were no false negative results; the false positive rate was determined and produced a value of <2%. Based on the data generated, the method demonstrated Assurance GDS for Cronobacter Tq II assay produced acceptable interlaboratory reproducibility data and statistical analysis. The Assurance GDS for Cronobacter Tq II method is suitable for the rapid qualitative detection of Cronobacter in infant formulas, infant cereals, ingredients, and environmental samples.

Inter-intra instrument comparison and standardization of a 10-color immunophenotyping for B and T cell non-Hodgkin lymphoma diagnosis and monitoring

Harmonization of flow cytometry protocols from instrument settings to antibody panel and reagents is highly encouraged for inter-laboratory data comparison in both research and clinical settings, especially for minimal residual disease monitoring evaluation in hematological diseases across centers. Here, we described inter-intra instrument comparison of two standardized 10-color staining dried tubes for B- and T-cell lymphoproliferative disorder diagnosis and monitoring on two different flow cytometers, a Beckman Coulter NaviosEx and a Beckman Coulter DxFlex. A total of 47 consecutive patients were enrolled, and 39 of them were evaluable for further studies. We show highly comparable results between the two cytometers for cell frequency and fluorescence intensity signals for both standardized 10-color staining dried tubes. For this latter, fluorescence of each antibody and subject was normalized on the mean value obtained from the entire study cohort thus reducing the effects of biological variability and allowing comparison between instruments with different detector sensitivity. In summary, dried tubes were confirmed as an optimal standardized diagnostic tool, especially when associated with EuroFlow standardized procedures by minimizing technical and biological variability. However, data analysis is still operator-dependent, and more efforts are needed to develop automated or semi-automated software for flow cytometry data analysis for diagnostic purposes.

Optimization of Aflatoxin B1-Lysine Analysis for Public Health Exposure Studies.

Aflatoxin B1 is a potent human carcinogen produced by several species of Aspergillus mainly found on nuts and maize. Exposures in parts of Africa, Latin America and Asia can be at multiples, sometimes orders of magnitude above tolerable daily levels. Although human exposure to aflatoxin can be estimated by analysis of the diet, only determination of the serum albumin aflatoxin adduct provides a health-relevant exposure measure. The lack of a reference serum limits interlaboratory method validation and data comparisons. In this study, we synthetically produced AFB1-dialdehyde and covalently coupled it to serum albumin in human serum. This synthetic produced aflatoxin-serum reference material was used in conjunction with isotopically labelled internal standards to evaluate sample digestion methods. This showed using sufficient Pronase in the digestion step was critical to ensure complete proteolytic digestion, which occurs within 4 h. Increasing the digestion temperature from 37 °C to 50 °C also provided a benefit to the overall analysis. In addition, the use of dried blood spots and Volumetric Absorptive Microsampling (VAMS) were investigated showing samples stored with VAMS produced equivalent results to serum samples.

Evaluation of the performance of modern X-ray fluorescence spectrometry systems for the forensic analysis of glass

Micro X-ray Fluorescence Spectrometry (µXRF) is a well-established technique for the elemental analysis of glass in forensic casework. The standard test method for the forensic analysis of glass using µXRF (ASTM E2926) provides recommendations for the number of replicate measurements that should be collected to characterize a known source and the criteria for the comparison between the known and questioned samples. However, these recommendations were based on interlaboratory data collected using µXRF instrumentation equipped with traditional lithium-doped silicon (SiLi) detectors. This interlaboratory study aimed to evaluate the performance of modern µXRF systems equipped with silicon drift detectors (SDDs) for the forensic comparison of glass. While the SDD-µXRF instruments resulted in improved precision and detection limits (1.4 µg·g−1–1386 µg·g−1) and excellent discrimination (>99 %) of different-source samples, the false exclusion rates for same-source samples were relatively high (>20 %). Two methods were evaluated to reduce the high false exclusion rates: increasing the number of fragments collected for the known source and modifying the recommended comparison criteria. To reduce the false exclusion rate to 5 % or less, a minimum of five known fragments were needed. Alternatively, modifying the recommended comparison criterion reduced the false exclusion rate from 23 % to 2 %, while maintaining low false inclusions (<1%). The findings in this study demonstrate the improved sensitivity and precision observed in glass measurements acquired with µXRF-SDD systems. However, these systems may require adjustments to sampling and the comparison criteria to minimize potential error rates in the forensic comparison of glass fragments.

Consistency plots: a simple graphical tool for investigating agreement in key comparisons

A simple graphical display is described for investigating agreement among interlaboratory data with reported uncertainties. The plot consists of a measure of agreement—the significance level of a test for significant pairwise difference, adjusted for multiple comparisons—plotted as an image in which significance is represented by colour or intensity. This provides an easily interpretable graphical presentation in which the degree of consensus can be judged and in which anomalies are easily visible. The construction of the plot is discussed, with attention to the choice of adjustment for the size of the data set in checking for anomalies. The advantage over visual inspection using error bars is discussed, and some examples from metrology comparisons are presented.

915P EMLI-ICC: An ensemble machine learning-based proteome and transcriptome integration algorithm for metastasis prediction and risk-stratification in intrahepatic cholangiocarcinoma

Intrahepatic cholangiocarcinoma (ICC) is the second-most-common primary liver cancer with increasing incidence and mortality worldwide. Although tumor metastasis is frequent even in ICC, improved robust strategies to identify patients at high risk for metastasis remains limited. Batch effects and different data types greatly decrease the predictive performances of signatures based on gene/protein expression profiles in inter-laboratory and different data type validation.

Simulation of the FDA nozzle benchmark: A lattice Boltzmann study

Background and objective: Contrary to flows in small intracranial vessels, many blood flow configurations such as those found in aortic vessels and aneurysms involve larger Reynolds numbers and, therefore, transitional or turbulent conditions. Dealing with such systems require both robust and efficient numerical methods.Methods: We assess here the performance of a lattice Boltzmann solver with full Hermite expansion of the equilibrium and central Hermite moments collision operator at higher Reynolds numbers, especially for under-resolved simulations. To that end the food and drug administration’s benchmark nozzle is considered at three different Reynolds numbers covering all regimes: (1) laminar at a Reynolds number of 500, (2) transitional at a Reynolds number of 3500, and (3) low-level turbulence at a Reynolds number of 6500.Results: The lattice Boltzmann results are compared with previously published inter-laboratory experimental data obtained by particle image velocimetry. Our results show good agreement with the experimental measurements throughout the nozzle, demonstrating the good performance of the solver even in under-resolved simulations.Conclusion: In this manner, fast but sufficiently accurate numerical predictions can be achieved for flow configurations of practical interest regarding medical applications.

Evaluation of the Thermo Scientific SureTectTM Listeria Species PCR Assay in a Broad Range of Foods and Selected Environmental Surfaces: Pre-Collaborative and Collaborative Study, First Action 2021.06.

BackgroundThe Thermo Scientific SureTect™ Listeria species PCR assay utilizes SolarisTM reagents for performing PCR for the rapid and specific detection of Listeria species in a broad range of foods and selected environmental surfaces.ObjectiveTo demonstrate reproducibility of the Thermo Scientific SureTect Listeria species PCR assay in a collaborative study using a challenging matrix, full-fat cottage cheese (25 g), to extend the scope of the method.MethodsIn the collaborative study, the candidate method was compared to the US Food and Drug Administration/Bacteriological Analytical Manual (FDA/BAM) Ch. 10 Listeria reference method. The candidate method used two PCR thermocyclers, the Applied Biosystems QuantStudio™ 5 Real-Time PCR instrument (QS5) and the Applied Biosystems 7500 Fast Real-Time PCR instrument (7500 Fast). The candidate method included its own confirmation procedure. Eighteen participants from 10 laboratories located within the United States and Europe were solicited for the collaborative study, with 12 participants submitting valid data. Statistical analysis was conducted according to the probability of detection (POD) statistical model. In addition, in order to extend the scope of the method, seven matrix studies were performed comparing the candidate method to the FDA/BAM reference method. One of these matrixes was also compared to the ISO 11290–1:2017 Microbiology of the food chain—Horizontal method for the detection and enumeration of Listeria monocytogenes and of Listeria spp.—Part 1: Detection method reference method.ResultsIn the collaborative study, the difference in laboratory results indicates equivalence between the candidate method and reference method for the matrix evaluated and the method demonstrated acceptable inter-laboratory reproducibility as determined in the collaborative evaluation. The two PCR instruments used in the study performed equivalently. All presumptive positives were confirmed via the alternative confirmation procedure. In the pre-collaborative studies, the results showed comparable performances between the candidate method and the reference method for all matrixes tested.ConclusionBased on the data generated, the method demonstrated acceptable inter-laboratory reproducibility data and statistical analysis.HighlightsDue to the COVID-19 pandemic, some participants had to be trained remotely. Additionally, 25 g full-fat cottage cheese is known to be a challenging matrix to test. No unusual cross-contamination, or false-positive/negative data was reported, highlighting the ease of use, reproducibility, and robustness of the candidate method.

Evaluation of the Thermo ScientificTM SureTectTMListeria monocytogenes PCR Assay in a Broad Range of Foods and Selected Environmental Surfaces: Pre-Collaborative and Collaborative Study, First Action 2021.05.

BackgroundThe Thermo Scientific™ SureTect™ Listeria monocytogenes PCR Assay uses Solaris reagents for performing PCR for the rapid and specific detection of Listeria monocytogenes in a broad range of foods and selected environmental surfaces.ObjectiveTo demonstrate reproducibility of the SureTect Listeria monocytogenes PCR Assay in a collaborative study using a challenging matrix, full-fat cottage cheese (25 g). To extend the scope of the method.MethodIn the collaborative study, the candidate method was compared to the United States Food and Drug Administration/Bacteriological Analytical Manual (FDA/BAM) Chapter 10 Listeria reference method. The candidate method used two PCR thermocyclers, the Applied Biosystems™ QuantStudio™ 5 Real-Time PCR instrument (QS5) and the Applied Biosystems 7500 Fast Real-Time PCR instrument (7500 Fast). Eighteen participants from 10 laboratories located within the United States and Europe were solicited for the collaborative study, with 12 participants submitting valid data. Three levels of contamination were evaluated for each matrix. Statistical analysis was conducted according to the probability of detection (POD) statistical model. In addition, to extend the scope, six matrix studies were performed comparing the candidate method to the FDA/BAM reference method. One of these matrixes was also compared to the ISO 11290–1:2017 Microbiology of the Food Chain—Horizontal Method for the Detection and Enumeration of Listeria monocytogenes and of Listeria spp.—Part 1: Detection Method Reference Method.ResultsIn the collaborative study, the difference in laboratory results indicates equivalence between the candidate method and reference method for the matrix evaluated, and the method demonstrated acceptable inter-laboratory reproducibility as determined in the collaborative evaluation. The two PCR instruments used in the study performed equivalently. All presumptive positives were confirmed via the alternative confirmation procedure. In the pre-collaborative studies, the results showed comparable performances between the candidate method and the reference method for all matrixes tested.ConclusionsBased on the data generated, the method demonstrated acceptable inter-laboratory reproducibility data and statistical analysis.HighlightsDue to the COVID-19 pandemic, some participants had to be trained remotely. Additionally, 25 g full-fat cottage cheese is known to be a challenging matrix to test. No unusual cross-contamination or false positive/negative data were reported, highlighting the ease of use, reproducibility, and robustness of the method.

Data accuracy and method validation of chemical soil properties in long-term experiments: Standard operating procedures for a non-certified soil laboratory in Latin America

Long-term agricultural experiments (LTE) are essential to detect cumulative treatment effects on soil properties and to design sustainable production systems. However, to ensure high quality of long-term soil data and their correct interpretation, several analytical issues regarding the accuracy and analytical laboratory bias need to be considered. This paper aims to (1) evaluate laboratory precision and trueness of analytical soil data for the evaluation of long-term trends in LTE and to (2) assess the limitations and challenges for non-certified soil laboratories that might compromise the quality of analytical soil data. A data set of internal reference soil materials (IRM) collected over 16 years and interlaboratory data from eleven years were analyzed to verify method precision, trueness, and the subsequent long-term dataset reliability for several soil quality parameters: organic carbon (SOC, determined either by wet or dry combustion), pH (water), extractable phosphorous (either Bray I or citric acid; Bray-P or citric acid-P), and exchangeable potassium (Kexch). Results showed that IRM used by the laboratory were homogenous in terms of physical and chemical composition and appropriate to confirm the precision of long-term soil survey data. The relative standard deviation for repeatability and reproducibility (RSDR) ranged from 1.5% for SOC by wet combustion to 9.5% for citric acid-P. HorRat values (the ratio of the estimated standard deviations of reproducibility and the repeatability found for individual analytical procedures) for all chemical soil properties were within the acceptable ranges of <2.0. Interlaboratory trials for soil pH and SOC showed tolerable standard Z-Scores under 2.0 (Z-Score, calculated from the difference between laboratory results and the assigned value divided by the standard deviation), verifying the trueness of data. The results of this study reinforce the validity of analytical soil data originating from the non-certified laboratory in Uruguay obtaining both precise and true soil quality data over a long period of time for most soil analytes. Nevertheless, the analytical flaws in LTE soil monitoring were revealed and can only be minimized in ongoing and future studies through the inclusion of certified reference material. These recommendations should guide future research activities in LTE studies on analytical data quality management as a requirement for long-term soil monitoring. Finally, the paper proposes a proficiency testing procedure for soil laboratories to achieve and maintain high analytical quality for LTE soil research.

Evaluation of the Thermo Scientific SureTect Salmonella Species PCR Assay in a Broad Range of Foods and Select Environmental Surfaces: Pre-Collaborative and Collaborative Study: First Action 2021.02.

BackgroundThe Thermo Scientific™ SureTect™ Salmonella species PCR Assay utilizes Solaris™ reagents for performing PCR for the rapid and specific detection of Salmonella species in a broad range of foods and select environmental surfaces.ObjectiveThe aims were to demonstrate the reproducibility of the Thermo Scientific SureTect Salmonella species PCR Assay in a collaborative study using a challenging matrix, cocoa powder, and to extend the scope of the method.MethodIn the collaborative study, the candidate method was compared to the US Food and Drug Administration (FDA) Bacteriological Analytical Manual (BAM) Chapter 5 Salmonella reference method. The candidate method used two PCR thermocyclers, the Applied Biosystems™ QuantStudio™ 5 Real-Time PCR instrument (QS5) and the Applied Biosystems 7500 Fast Real-Time PCR instrument (7500 Fast). Fourteen participants from nine laboratories located within the United States and Europe were solicited for the collaborative study, with 12 participants submitting valid data. Three levels of contamination were evaluated for each matrix. Statistical analysis was conducted according to the probability of detection statistical model. In addition, 11 matrix studies were performed comparing the candidate method to the FDA/BAM Chapter 5 or US Department of Agriculture, Food Safety and Inspection Service, Microbiology Laboratory Guidebook 4.10 Isolation and Identification of Salmonella from Meat, Poultry, Pasteurized Egg, and Siluriformes (Fish) Products and Carcass and Environmental Sponges reference method. Nine of these matrices were also compared to the EN ISO 6579–1:2017/Amd.1:2020(E) Microbiology of the food chain—Horizontal method for the detection, enumeration and serotyping of Salmonella—Part 1: Detection of Salmonella spp.—AMENDMENT 1: Broader range of incubation temperatures, amendment to the status of Annex D, and correction of the composition of MSRV and SC reference method.ResultsIn the collaborative study, the difference in laboratory results indicates equivalence between the candidate method and reference method for the matrix evaluated, and the method demonstrated acceptable interlaboratory reproducibility as determined in the collaborative evaluation. False-positive and false-negative rates were determined for the matrix and produced values of <2%. The two PCR thermocyclers (QS5, 7500 Fast) performed equivalently. There were no result differences between candidate method confirmations and reference method confirmations. In the pre-collaborative matrix extension, the results from the matrix studies showed a comparable performance between the candidate method and the tested reference methods.ConclusionsBased on the data generated, the method demonstrated acceptable interlaboratory reproducibility data and statistical analysis.HighlightsDue to the COVID-19 pandemic, some participants had to be trained remotely. Additionally, 375 g cocoa powder is known to be a challenging matrix for PCR methods. No unusual cross-contamination or false-positive/negative was reported, highlighting the ease of use, reproducibility, and robustness of the method.

Construction of an in-house allelic ladder for Canine Genotypes™ Panel 2.1 Kit.

Dogs (Canis lupus familiaris) are among the most common companion animals in the Republic of Korea. Recently, there have been many criminal cases of dog cruelty, injury, and theft, among others. This has increased the importance of dog-related biological evidence at crime scenes. The National Forensic Service of the Republic of Korea conducts short tandem repeat (STR) analysis using the Thermo Scientific Canine Genotypes™ Panel 2.1 Kit (Canine Kit) to identify individual dogs through forensic analysis. The Canine Kit was developed as a forensic STR kit for the identification of individual dogs. However, an allelic ladder was neither developed nor included in the commercial kit, leaving an issue of accurate genotyping. Primer details for the 18 markers used in the Canine Kit are proprietary information, and thus, unavailable to end-users. In this study, an allelic ladder was constructed with 160 fragments by combining 158 fragments of STR alleles obtained by nested PCR and two fragments artificially obtained from the sex-determination marker. By including the new allelic ladder in analysis of samples amplified with the Canine Kit, the accuracy and reliability of data analysis were improved. Application of this allelic ladder would be helpful for interlaboratory data sharing and standardization of canine genotype databases.