Search Results (99)

Newborn screening in England is a national program with laboratories adhering to common screening algorithms. Until recently, screening for inherited metabolic disorders was provided by ten laboratories using laboratory-developed tests (LDTs) and three using commercial assays: harmonization of results proved challenging. Introduction of hereditary tyrosinemia type 1 screening meant LDTs required modification to include the measurement of succinylacetone, and subsequent re-validation. This provided an opportunity to implement a single commercial reagent kit in all laboratories. It was anticipated that this would improve analytical performance and harmonization. This study aimed to determine whether these goals were achieved. Verification across the 13 laboratories revealed that the commercial kit reduced inter-laboratory variation for all analytes demonstrating improved harmonization. However, this was achieved by applying instrument-specific correction factors to all analytes, the magnitude of which were significant, indicating a lack of standardization. Performance of succinylacetone was limited by instrument-dependent background interference from the methionine stable isotope label, underscoring the need to establish evidence-based screening cut-off values (COV) rather than adopting published thresholds. This study emphasizes the need for traceable reference materials to improve laboratory quality and the value of screening outcome data. Full article

Cereulide is a heat-stable cyclic depsipeptide toxin produced by Bacillus cereus and is responsible for foodborne emetic syndrome. Recent reports of Bacillus cereus contamination and cereulide occurrence in infant formula have raised increasing food safety concerns. Due to the immature immune and metabolic systems of infants, exposure to cereulide through contaminated formula may lead to potential health risks. However, direct application of existing cereulide analytical methods to infant formula remains challenging because of the unique processing technologies, encapsulated nutrients, and variable matrix composition of this product category, which may hinder toxin release and cause significant matrix interference. In practical analysis, inter-laboratory comparisons revealed that existing methods exhibited relatively large deviations and insufficient sensitivity, making them not specifically optimized for infant formula matrices. The present study was motivated by the need for a matrix-specific, sensitive, and reliable analytical method for cereulide determination in infant formula. In this study, a method based on solid-phase extraction coupled with ultra-performance liquid chromatography–tandem mass spectrometry (SPE–UPLC–MS/MS) was developed and validated. To improve the applicability of cereulide analysis to infant formula, this method incorporates a hydration-assisted extraction step tailored to infant formula, which increased the detected cereulide response by approximately fourfold, together with optimized SPE clean-up and improved chromatographic conditions to reduce matrix effects and enhance quantitative reproducibility. The method showed good linearity (0.1–10 μg·L⁻¹, R² > 0.999), low values for limit of detection (LOD) (0.03 μg·kg⁻¹) and limit of quantification (LOQ) (0.1 μg·kg⁻¹), and acceptable recoveries (94.4–110.3%) with RSDs below 3.7%. The developed method was successfully applied to commercial infant formula samples, and cereulide-positive samples were identified. This method provides a reliable analytical tool for the monitoring of cereulide in infant formula and contributes to improved food safety surveillance and exposure risk assessment. Full article

The evaluation of D- and L-lactic acid production by lactic acid bacteria is of critical importance, particularly for strains intended for use in infant and young child foods. Additionally, compliance with relevant regulatory standards necessitates the detection of D-lactic acid. Additionally, regulatory requirements exist in this regard. This study aimed to develop and validate a method for simultaneously measuring D-lactic acid and L-lactic acid produced by lactic acid bacteria. The method validation of the HPLC analysis was performed in terms of accuracy, precision, specificity, limit of quantification (LOQ), linearity, range, and robustness, and the measurement uncertainty was also evaluated. The method demonstrated a limit of detection (LOD) of 0.25 µg/mL and LOQ of 0.8 μg/mL for D-/L-lactic acid. For six validated bacterial strains, mean recoveries ranged from 93.50% to 105.37%, with intra-assay relative standard deviations (RSD) of 0.90–2.64% and inter-laboratory RSD of 2.56–10.16%. Excellent linearity, accuracy, and precision were observed across the concentration range of 0.8–200.0 μg/mL. Results confirmed no interference from culture media batch variations, and sample stability was maintained for 48 h. Additionally, relative expanded uncertainties were determined as 10.48% and 7.64%. The developed method was suitable for the identification and quantification of D- and L-lactic acid in lactic acid bacteria fermentation broth samples. This method was applicable for assessing the production of D-/L-lactic acid by food cultures. Full article

Urinary epidermal growth factor (uEGF) and clusterin (uCLU) are emerging biomarkers in chronic kidney disease (CKD), but rigorous analytical validation is required before clinical implementation. We evaluated intra-individual variability and long-term storage stability of uEGF and uCLU in CKD. In the prospective, multicenter UVALID study, 60 adults with CKD stages 2–4 underwent urine sampling at three visits over 8 weeks. First-morning and 24-h urine samples were collected to assess intra-individual variability over 24 h, 3 days and 8 weeks. Biomarkers were measured in duplicate by ELISA and normalized to urinary creatinine (/Cr). Inter-laboratory performance was assessed using quality control samples. Stability after 12 and 15 months of storage at −20 °C and −80 °C and the influence of pH were evaluated. Over 24 h, 3 days, and 8 weeks, uEGF/Cr demonstrated low variability and remained stable after long-term storage at both temperatures. In contrast, uCLU/Cr showed greater variability and pronounced instability at −20 °C, whereas stability was preserved at −80 °C. Samples with pH > 6 partially preserved uCLU stability at −20 °C. Inter-laboratory reproducibility was acceptable for uEGF but suboptimal for uCLU at low concentrations. Thus, uEGF showed robust analytical performance, supporting its potential clinical applicability in CKD, whereas uCLU exhibited important analytical and pre-analytical limitations, warranting further assay optimization. These findings underscore the need for rigorous validation to facilitate biomarker implementation in clinical practice. Full article

Perovskite solar cells (PSCs) have quickly achieved certified energy conversion efficiency reaching a certified record of 27.3% for single-junction cells, while having a low mass, thin-film form factor and high specific power, which are attractive for space energy systems. However, their long-term reliability in extraterrestrial environments is not adequately ensured by terrestrial qualification routes, and standardized space-related test protocols remain insufficiently developed. This review critically summarizes the current understanding of the degradation of PSCs under the influence of key environmental factors in space—ionizing and non-ionizing radiation, thermal vacuum exposure and thermal cycling, and ultraviolet radiation AM0, as well as atmospheric oxygen in low orbits. The central task of the work is to develop and justify the need to create specialized PSCs test protocols for space applications, since existing ground standards do not reflect the multifactorial nature and extreme orbital loads. It has been shown that thermal vacuum accelerates ion migration, interphase reactions, and degassing, while AM0 UV and atomic oxygen introduce additional photochemical and oxidative mechanisms of destruction; at the same time, stressors often act synergistically and are not detected by single-factor tests. Next, the limitations of the current IEC and ISOS are discussed and an approach to their expansion is formulated through the ISOS-T-Space and ISOS-LC-Space protocols, which integrate high vacuum, AM0 lighting, extended temperature ranges and controlled particle irradiation. It is concluded that the development and interlaboratory validation of such space-oriented protocols is a key condition for the correct qualification of PSCs and targeted optimization of materials and interfaces to meet the requirements of space energy. Full article

Fabrazyme (agalsidase beta) is a therapeutic enzyme whose clinical efficacy is contingent upon its complex N-glycosylation patterns. Nevertheless, comprehensive glycosylation profiling remains challenging due to high site-specific heterogeneity. To address this, three orthogonal liquid chromatography–mass spectrometry (LC-MS) approaches were employed: (1) released N-glycan analysis with fluorescence detection and MS annotation, (2) site-specific glycopeptide mapping, and (3) intact protein MS. The released glycan profiling method was validated for reproducibility, intermediate precision, and inter-laboratory transferability, thereby enabling reliable separation and quantification of neutral, phosphorylated, and sialylated species. Glycopeptide mapping revealed distinct site-specific distributions: N108 was found to predominantly carry sialylated complex glycans; N161 was enriched in phosphorylated oligomannose structures; and N184 displayed the highest heterogeneity, including bisphosphorylated and sialylated glycans. Intact protein analysis was performed on both intact and desialylated Fabrazyme, thereby enabling confirmation of glycan assignments. Desialylation reduced spectral complexity, thereby facilitating accurate mass matching with a combinatorial library generated from glycopeptide-level data. The complementary use of these three analytical levels provides a comprehensive view of Fabrazyme glycosylation, offering a robust reference for quality control and biosimilar development. Full article

Ips typographus is one of the most destructive bark beetles affecting conifer forests in Europe, where climatic disturbances and the movement of infested wood can rapidly shift populations from endemic levels to severe outbreaks. Early detection through border inspections and forest monitoring is essential to prevent new introductions and limit the spread of established populations. Here, we developed and validated a probe-based TaqMan qPCR assay, targeting the mitochondrial COI barcode region, for the rapid and species-specific detection of I. typographus from both insect material and environmental DNA recovered from frass and exit-hole wood chips. Validation followed EPPO PM7/98(5) guidelines, assessing analytical specificity, sensitivity, repeatability, reproducibility, and inter-laboratory transferability. High analytical specificity was demonstrated against a broad panel of non-target species, and reliable amplification was obtained across different tested matrices. The method showed strong analytical sensitivity, with limits of detection of 0.32 pg/µL for adult-derived DNA and 1.6 pg/µL for artificial frass. Repeatability, reproducibility, and inter-laboratory blind testing further confirmed the diagnostic reliability of the method. This validated qPCR assay provides a rapid and sensitive molecular tool for the early detection of I. typographus, supporting border inspection and phytosanitary diagnostic laboratories in forest biosecurity activities. Full article

Background/Objectives: Breast cancer recurrence risk stratification has relied on gene expression tests that are limited by long turnaround times and consumption of valuable tissue. Artificial intelligence (AI) utilizing digital pathology images elucidates novel morphological biomarkers with strong prognostic associations, but the use of such AI models requires a modified analytical validation approach. Here, we report analytical validation of a novel breast cancer prognostic test. Methods: Ataraxis Breast RISK (ATX) uses a survival analysis model based upon features from a pan-cancer foundation model. This model extracts morphological features (biomarkers) from H&E-stained slides. These features are combined with clinical variables, and the test outputs a calibrated recurrence risk score. We validated ATX across five axes: intra-operator repeatability, inter-operator reproducibility, limit of blank, limit of detection and inter-laboratory reproducibility. Additionally, we assessed robustness to clinicopathologic data perturbations and conducted a clinical validation bridging study. Experiments were performed in CLIA-certified laboratories. Results: Intra-operator repeatability yielded an intraclass correlation coefficient (ICC) of 0.99 with 100% risk category agreement. Inter-operator reproducibility was concordant (ICC 0.99, 100% agreement). Inter-laboratory reproducibility across multiple scanners showed an ICC of 0.97 with 94.7% agreement. Under simulated clinicopathologic data perturbation, ATX maintained an average C-index of 0.62 with 90.0% agreement. The bridging study confirmed that the performance of the CLIA version was comparable to the prior clinical validation version (C-index 0.63 vs. 0.62). Conclusions: ATX met all predefined analytical acceptance criteria. These results support the analytical readiness of ATX use in clinical testing. Full article

The Epi2SensA method is a method similar to the validated EpiSensA assay for assessing the skin sensitization potential of chemicals. The Epi2SensA protocol includes adaptation (changes to exposure conditions and the controls) for using an alternative reconstructed human epidermis (RhE) model, the EpiDerm™ model. The interlaboratory validation study evaluated the reliability and predictive capacity of Epi2SensA according to OECD Performance Standards. Four laboratories (Mattek, Now Part of Sartorius, Eurofins Munich, Burleson Research Technologies, Inc., and Food and Drug Safety Center) conducted blinded testing of 20 coded reference substances representing various chemical categories and sensitization potencies. Statistical analysis using modified acceptance criteria (a 60% cell viability threshold) and a modified prediction model (requiring at least two positive gene markers) demonstrated substantially improved performance compared to the original EpiSensA criteria. The between-laboratory reproducibility (BLR) was 85%, the average within-laboratory reproducibility (WLR) was 83.3%, and the average predictivity parameters were 88.1% for sensitivity, 88.9% for specificity, and 88.3% for accuracy. Epi2SensA achieved performance metrics comparable to the validated reference method (EpiSensA), supporting regulatory acceptance of the Epi2SensA assay using the EpiDerm™ model (Mattek Corporation, Now Part of Sartorius, Ashland, MA, USA) as an alternative RhE source for OECD TG 442D skin sensitization testing. Full article

Background/Objectives: Biorelevant in vitro dissolution testing is used increasingly to predict complex mechanisms in the gastrointestinal (GI) tract that determine oral bioavailability. However, the limited use of non-compendial systems is driven by the lack of widely accepted, standardized validation frameworks. This ongoing gap continues to restrict their adoption relative to United States Pharmacopeia (USP) apparatus. While the physiological relevance and biopredictive capabilities of the tiny-TIM and TIM-1 in vitro GI models have been demonstrated in previous studies, their inter-laboratory reproducibility has not been systematically established. Therefore, this international ring study evaluates the reproducibility of in vitro simulations of GI transit and absorption of paracetamol in fasted- and fed-state conditions in tiny-TIM and TIM-1. Methods: Three laboratories used TIM-1 and five used tiny-TIM to simulate oral administration of a 500 mg paracetamol solution to a healthy adult. Paracetamol solution was selected as a well-characterized and widely available BCS I compound to minimize formulation and solubility effects and focus on system performance, enabling the generation of a generic validation dataset for the reproducibility of TIM experiments. Results: Paracetamol bioaccessibility profiles were repeatable and reproducible (all pairwise f₂ > 50). Maximum differences in total bioaccessible paracetamol were 0.9% (TIM-1) and 2.8% (tiny-TIM) within laboratories and 3.4 and 5.9% between laboratories. Inter-lab variability at individual time points remained <4.0% (fasted) and 5.2% (fed). Both TIM models produced biopredictive metrics, correctly predicting no food effect on total paracetamol bioaccessibility and capturing delayed t_max. Gastric and intestinal environments showed repeatable pH, temperature, and GI transit characteristics, with fluctuations across transit stages that mirrored reported in vivo patterns. Conclusions: These results demonstrate that TIM systems can reproducibly simulate gastrointestinal conditions across laboratories and generate consistent measurements of drug product performance, despite the complexity of the dynamic processes involved. While this evaluation involving a single BCS I drug solution should not be directly extrapolated to experiments with poorly soluble compounds or different formulations, it supports the use of TIM systems as robust in vitro models in drug product development. This study provides a standardized, inter-laboratory, baseline performance dataset to support regulatory submissions incorporating TIM data and enable more confident interpretation of TIM experiments. Full article

To address the issues of operational complexity, long duration association, and reliance on specialized equipment with existing detection methods for Vibrio parahaemolyticus, this study established a rapid detection method for V. parahaemolyticus in exported aquatic products based on the domestically developed Enzymatic Recombinase Amplification (ERA) technology. To target the thermolabile hemolysin gene (tlh) and the iron-regulated virulence regulatory protein gene (irgB) of V. parahaemolyticus, highly specific ERA primers and probes were designed and screened. Two detection platforms, a colorimetric method and a fluorescent method, were developed. Method validation results showed that this detection system achieved specific amplification for all 30 tested V. parahaemolyticus strains, with no cross-reactivity observed with 30 other common foodborne pathogenic bacteria. The detection sensitivity for both the fluorescent and colorimetric methods reached 10⁻¹ ng/μL, with a minimum detection limit of 10 CFU/25 g for artificially contaminated samples. The entire detection process, including sample preparation, requires only approximately 20 min—significantly faster than traditional culture (24–72 h) or even conventional PCR methods. Collaborative validation across five independent laboratories confirmed excellent reproducibility, with inter-laboratory agreement yielding a Kappa coefficient of 0.98. The ERA method operates at a low, constant temperature (37–39 °C), eliminating the need for thermal cyclers. When combined with portable isothermal amplification devices and visual (colorimetric) readout, it offers a distinct advantage in terms of speed, cost-effectiveness, and suitability for resource-limited or field settings compared to existing PCR-based or culture-based platforms. This method is simple to operate, rapid, sensitive, and highly suitable for on-site application, providing a reliable and practical technical solution for the rapid screening and risk monitoring of V. parahaemolyticus in exported aquatic products. Full article

This study details the validation of a novel microbead beating extraction (MBE) technique for the analysis of polycyclic aromatic compounds (PACs) in sediments. The method’s performance was evaluated against international analytical validation criteria, including trueness, precision, measurement uncertainty and robustness. Limits of detection and quantitation were consistently low (≤6.5 and 21 ng g⁻¹, respectively), trueness for the majority of analytes fell within accepted performance criteria, and repeatability values for most analytes were generally <10%. Analytical data confirm the method’s reliability, with more than 80% of certified analytes in two certified reference materials (CRMs) meeting the satisfactory z-score (∣z∣ ≤ 2.0). Furthermore, the method’s inter-laboratory repeatability, as measured by HorRat values, fell within the range recommended by the Association for Official Analytical Chemist for most analytes, and combined measurement uncertainties showed no statistical difference from the certified uncertainties of the CRMs. Incorporating an in situ cleanup step enabled the MBE method to substantially reduce extraction times (<15 min) and reduces solvent consumption by ~60% compared with conventional pressurize fluid extraction while maintaining good quality data objectives. By meeting or exceeding well-established metrics for good laboratory performance, the MBE method demonstrates reliability, efficiency, and a greener approach for the routine analysis of PACs in sediments. Full article

Microplastics (MPs) are emerging pollutants with widespread global distribution, continuously accumulating in soils and posing risks of cross-media pollution. Current soil MP detection methods lack unified standards, suffering from high inter-laboratory variability and cost, which become key bottlenecks limiting data comparability and global microplastics pollution control. Here, we systematically reviewed soil MPs studies (2020–2024) and based on stepwise verification, we established a standardized, reproducible detection method: soil samples were dried at 80 °C for 12 h; density separation was performed in Erlenmeyer flasks with decantation, 10 s glass rod stirring, and 12 h settling, repeated five times; digestion was conducted using a 1:2 volume ratio of H₂O₂ to supernatant at 80 °C for 8 h; and MPs were quantified via stereo-microscopy combined with ImageJ. It should be noted that the use of NaCl limits the recovery of high-density polymers (e.g., PVC, PET), and the minimum detectable particle size is approximately 127 µm. The method was validated in sandy, loam, and clay soils, achieving an average recovery rate of 96.4%, with a processing time of 68 h and a cost of USD 9.77 per sample. In contrast to previous fragmented, non-standardized protocols, this workflow synergistically optimizes high recovery efficiency, cost-effectiveness, and broad applicability, offering a low-cost, efficient, and widely applicable approach for soil MPs monitoring, supporting data comparability across studies and contributing to global pollution assessment and the United Nations 2030 Sustainable Development Goals. Full article

International comparisons play a critical role in ensuring precision, accuracy, consistency, and trust in the global metrology system. The Comité International des Poids et Mesures (CIPM) established the Mutual Recognition Arrangement (MRA) in 1999 to facilitate global trade. This paper gives an overview of the critical role of international comparisons to National Metrology Institutes (NMIs), industrial calibration laboratories, and research laboratories in fostering global measurement equivalence and the CIPM MRA. NMIs rely on Key and Supplementary Comparisons to ensure the mutual recognition of calibration and reference material certificates, vital for global trade and regulatory compliance. Industrial calibration laboratories participate in inter-laboratory or international comparisons to validate their calibration and measurement capability (CMC) and balance their risk management. Research laboratories push the frontiers of measurement science and validate their measurement result via international comparisons. Through some examples of comparisons, the paper illustrates how measurement result discrepancies uncovered in comparisons drive technical improvements, uncertainty component identification, and measurement technique refinement. International comparisons enhance scientific credibility, build public trust, support industrial innovation, and drive evolution in measurement science. As technological demands grow, fostering broader participation in international comparisons by various metrology and research laboratories remains crucial to maintain a robust and reliable global metrology system. Full article

Feline leishmaniosis (FeL) is still considered an emerging and neglected disease. Cats, once considered accidental hosts, are now recognized as adjunctive reservoirs of the disease, especially in areas where canine (CanL) and human (HumL) leishmaniosis are widespread. Although often asymptomatic, infected cats could contribute to the transmission cycle of the parasite. Recent studies in Campania (Italy) have found a significant prevalence of feline infection, indicating the need to implement diagnostic and surveillance protocols to prevent the spread of the disease. The aim of the study was to outline the current scenario by studying the prevalence of FeL in Campania to identifying the potential zoonotic risk and in addition to validate the Immunofluorescence Antibody Test (IFAT) method for the diagnosis of leishmaniosis in cats. The study involved initially 702 cats; for each cat, a clinical record was compiled, including identification data, anamnesis, and clinical findings. Due to incomplete information, statistical analysis was performed only on a subset of 601 cats. A blood sample was collected to obtain serum/plasma specimens. When feasible, a lymph node fine-needle aspiration was performed. The observed seroprevalence rate was 32.1% (193/601), with a higher seroprevalence in outdoor cats and the presence of asymptomatic seropositive animals (28.0%;54/193), suggesting that felines may act as silent reservoirs of Leishmania infantum. An excellent result was obtained for the validation and standardization of the analytical IFAT method for the diagnosis of feline leishmaniasis; therefore, an inter-laboratory test has been carried out to establish the dilution cut-off at ≥1:80 as compatible with infection. Furthermore, a xenodiagnosis examination was conducted on a cat that was infected to more accurately evaluate the possibility of asymptomatic cats acting as carriers of the infection; however, this test resulted negative. Full article