Volume 64, Issue 2

With the advent of flow cytometry and dedicated fluorescent dyes in the 1980s, automated reticulocyte enumeration has effectively superseded conventional manual microscopy, dramatically boosting throughput and precision while, through metrics including the immature reticulocyte fraction (IRF) and reticulocyte production index (RPI), delivering previously unattainable diagnostic resolution for anemia phenotyping, longitudinal tracking of marrow erythropoiesis, and evaluation of functional iron deficiency. Yet appreciable heterogeneity in analytical principles, reagent chemistry, and calibration frameworks persists across platforms, constraining result comparability and uniform interpretation across systems. The International Council for Standardization in Haematology (ICSH) is advancing commutable reference materials built on stable cell lines or synthetic microspheres, together with globally harmonized reference intervals, to achieve interoperable performance among instruments. Concurrently, the capital expense and servicing requirements of high-end fully automated analyzers continue to be a major impediment to clinical access to automated reticulocyte testing. This narrative review focuses on the “standardization and clinical accessibility of automated reticulocyte counting”, systematically elaborating the core technical principles and characteristics of mainstream instruments, analyzing advances and challenges in standardization, and envisioning its application prospects in global clinical practice.

EU laws are usually evaluated after 10 years; in the case of the IVDR, the targeted evaluation was brought forward and stakeholders in laboratory diagnostics were invited to comment. The Association of the Scientific Medical Societies in Germany (AWMF; Arbeitsgemeinschaft der Wissenschaftlichen Medizinischen Fachgesellschaften e.V.), recognises a relevant need to amend the IVDR with regard to in-vitro -diagnostic medical devices that are manufactured and used in healthcare facilities but are not placed on the EU market. Inadequate overregulation is recognised and should be corrected as part of the generally required reduction of bureaucracy.

The concept of value, defined as health outcomes achieved per monetary unit spent, has profoundly reshaped modern healthcare delivery. While Value-Based Healthcare models have permeated many clinical disciplines, laboratory medicine has been slow to integrate this paradigm shift. In this opinion paper, we argue for a strategic repositioning of clinical laboratories as core enablers of value in healthcare systems. Laboratory diagnostics, long considered ancillary, should be reframed as pivotal tools that support outcome-based, cost-effective decision-making. We explore how laboratory parameters contribute to clinical value through predictive accuracy, diagnostic specificity, and operational appropriateness – factors that directly influence patient outcomes and resource allocation. Examples such as vitamin D testing, albumin as a biomarker of biological age, and NT-proBNP in heart failure demonstrate the potential and pitfalls of volume-driven laboratory utilization. Beyond technical excellence, we emphasize the importance of interpretive collaboration, health literacy, and ethical stewardship of diagnostic resources. Structural challenges, including commoditization, delocalization via point-of-care testing, and the limited use of patient-reported outcomes in laboratory settings, are critically examined. Finally, we highlight emerging policy frameworks across Europe that align reimbursement models with measurable outcomes, advocating for the integration of laboratories in clinical governance and value-based procurement. In this renewed perspective, laboratories are not merely data providers but agents of personalized, sustainable, and patient-centered care.

Currently, there is no accepted standard practice for determining the frequency of EQA challenge. The challenge frequency has evolved based on history and local requirements. However, EQA frequency should be based on identifying patient risks caused by poorly performing laboratories, methods, or processes in any phase of the total testing cycle. The role of IQC is to ensure result consistency from day to day and to halt reporting of results if there is an analytical failure. Historically, both activities have been based on synthetic control material. With the development of patient-based approaches to IQC and EQA, it is possible to continuously monitor analytical systems using the same patient parameter, usually the mean or median. These techniques can provide laboratories with additional information to reduce patient risk. There are limitations of Patient-Based Quality Assurance (PBQA), fundamentally the lack of middleware and connection of the analyzers to the EQA provider. It cannot be used to monitor the success of harmonization/standardization of assays to a reference measurement procedure. But the use of patient-based approaches offers an opportunity to reconsider how EQA can be undertaken and the relationship between IQC and EQA. If PBRTQC and PBQA could be implemented to provide daily peer group comparisons, then method-specific bias could be identified quickly by a laboratory. If this could be supplemented with a commutable, reference value assigned EQA program, then monitoring harmonization/standardization of assays to a reference measurement procedure could be achieved.

Objectives Oral squamous cell carcinoma (OSCC) is the most prevalent form of squamous cell carcinomas of the head and neck (SCCHN), accounting for over 90 % of all oral cavity malignancies (approximately 275,000 new cases are worldwide diagnosed annually). Early-stage oral squamous cell carcinoma (T1 and T2) has a 5-year survival rate of up to 80 %. Survival rates decrease to 20–30 % at later stages (T3-T4). Each year, there are between 275,000 and 300,000 new cases of OSCC, and over 150,000 deaths worldwide. OSCC are usually non-hereditary tumors, although familial epidemiology has been recently reported. Methods From 2022 to 2024, we enrolled 56 patients from the complex structure of maxillofacial surgery and ORL, National Cancer Institute – IRCCS – Fondazione G. Pascale. The individuals enrolled underwent molecular testing via a multigene panel of 56 genes related to cancer predisposition customized in our laboratory. The panel included BRCA1 and BRCA2 . Results We identified a total of 7 pathogenic mutations annotated in clinical databases as ClinVar, in BRCA2 (two different variants), BRCA1 , MUTYH , BRIP1 , FANCM and FANCC genes (approximately 12.5 % of our patients). The results show a frequent predisposition to head and neck tumors similar to or even greater than that observed in other types of neoplasia, such as breast and ovarian cancers or colon cancer), with a predisposition of approximately 10 %. Conclusions Our results confirm that, similarly to other more studied tumors, predictive genomic medicine can play a crucial role in the early identification of germline mutations in head and neck cancers. This approach should be considered for the early detection of OSCC particularly for individuals at increased risk, e.g., those with a family history of the disease, who may also be candidates for targeted molecular therapies based on their genetic profile.

Objectives To develop a proactive risk assessment model for human papillomavirus (HPV) genotyping testing by integrating Failure Mode and Effects Analysis (FMEA) with quality indicators (QIs), ensuring compliance with ISO 15189:2022 and improving diagnostic accuracy. Methods A multidisciplinary team designed and performed detailed FMEA across pre-analytical, analytical, and post-analytical phases of HPV genotyping testing. To improve objectivity, we integrated Sigma metrics into the FMEA framework through a molecular diagnostics-specific model of QIs (MQI). The FMEA model systematically identified testing phases, potential failure modes, their effects, root causes, and existing controls. Risk was quantified using Severity, Occurrence (from 1-year QI data), and Detection scores (1–5 scale). Risk Priority Numbers (RPNs) were calculated (Severity × Occurrence × Detection) to prioritize failure modes, with mandatory interventions implemented for high-risk items (RPN≥40). Results Five high-risk failure modes (e.g., sample misidentification, data analysis errors) were identified and successfully mitigated to acceptable levels (RPN<40) through process optimization and standardization, achieving RPN reductions of 20–80 %. We established a molecular diagnostics-specific MQI, comprising 14 pre-analytical, 25 analytical, and three post-analytical phase QIs. QI-based risk assessment of 35 evaluable QIs for HPV genotyping testing revealed one high-risk QI (“Incorrect results due to information system failures”) and three medium-risk QIs, all of which were addressed through corrective actions. Conclusions This study developed an integrated FMEA-QI model for HPV genotyping testing, establishing both a traditional FMEA framework and a molecular diagnostics-specific MQI. The combined approach improves risk assessment objectivity and enables multidimensional analysis compared to conventional methods.

Objectives Transporting blood samples with pneumatic tube systems (PTSs) is routine in many hospitals. The influence of PTS transport on cell-free DNA (cfDNA) and circulating tumor DNA (ctDNA) analyses has not been comprehensively tested. Methods Paired blood samples from non-patient donors and cancer patients were transported manually or by PTS at two Danish hospitals using different PTS. A g-logger evaluated the forces applied to samples in each transport method. The concentration of short and long cfDNA fragments and ctDNA markers was analyzed using droplet digital PCR (ddPCR) and compared according to transportation. Different blood collection tubes (BCTs) were also investigated. Results All transportation methods subject samples to significantly different forces. The cfDNA concentration was slightly higher in samples transported with the Sumetzberger PTS than manually. There were no differences in cfDNA, whether using EDTA or Roche BCTs transported using Tempus600 or manually. Colorectal cancer patient samples showed insignificant changes in ratio of fragment lengths when transported by PTS compared to manual transport. In metastatic cancer patients, only a minor change in long/short fragment ratio was observed in PTS-transported samples. No discrepancy in ctDNA concentration/fraction was observed in cancer patient samples. Higher yields of cfDNA were obtained from EDTA samples, but no other differences were observed among the tested BCTs. Conclusions Although PTS transport introduced minor variations in cfDNA concentrations, these did not impact ctDNA interpretation. However, as PTSs vary in design, their pre-analytical effects should be carefully assessed, particularly as ctDNA analysis becomes more widely implemented in clinical practice.

Objectives Regulatory guidelines recommend non-parametric Passing–Bablok regression for evaluating the agreement between two measurement methods in laboratory settings. However, concluding for the agreement if the 95 % CI of the slope and of the intercept include 1 and 0, respectively is incorrect since the agreement assessment must focus on a null hypothesis of not equivalence and an alternative hypothesis of equivalence. Methods We exhaustively simulated appropriate structural models with several values of slope, intercept and measurement error by keeping equal variances and means of the two methods. We calculated the slope and intercept bias of four regressions: non-parametric Passing–Bablok, Theil, Ordinary Least Squares and Deming. In addition, we calculated the percentages of the agreement according to the not shareable Passing–Bablok suggestion. Furthermore, we calculated the percentages of the 95 % CI of the slope and of the intercept included within sensible equivalence thresholds for assessing the agreement. Results Passing–Bablok procedure gives unbiased estimates, a little more and less biased than those from Deming’s regression. The percentages of rejecting the hypothesis of no-agreement, according to the wrong Passing–Bablok’s approach are correctly near to 0.05 Type I error under the agreement and also for 0.990≤slopes≤1.005. However, they are too low for slopes >1.05 and <0.950. Conclusions The Passing–Bablok 95 % CIs are too wide for being included in sensible agreement thresholds according to a population equivalence model and, finally, this approach cannot be considered under the best agreement model of the individual equivalence.

Objectives The International Organisation for Standardisation (ISO) 15189 standard provides the requirements for quality and competence in medical laboratories. It aims to ensure that laboratories deliver accurate and reliable results for patient care. The first version of the same was published in 2003, followed by editions in 2007, 2012 and 2022. The objective of this article is to analyze the ethical commitments and requirements assumed to achieve reliable results in laboratory medicine incorporated in the ISO standard. Methods We compare the concepts and changes related to ethics introduced over time in the ISO 15189 versions. We also present, a comparative study about the evolution of ethical concepts in the clinical laboratory and how these changes have been reflected in ISO 15189 standards through 2003 to 2022. Results After a thorough comparative analysis, we observe that the ISO 15189:2022 version has strengthened the requirements for several key concepts of Ethics such as impartiality and confidentiality, and addressed the issues of laboratory users with regards to autonomy, beneficence and justice although these terminologies are not explicitly mentioned. However, several other key words and concepts have not been addressed with adequate emphasis. Conclusions We recommend that several key concepts related to ethics may need to be included in the future revisions, especially those related to new and upcoming technologies being used in laboratory diagnostics and research.

Objectives Clinical laboratory operations consume large amounts of electricity and water and generate considerable quantities of solid waste. In this study, we quantified and compared the estimated solid waste produced from performing one million basic metabolic panels (BMPs) across five major high-throughput chemistry analyzer systems commonly used in the United States. Additionally, we estimated the carbon emissions associated with the electricity and water usage of each analyzer to better understand their environmental impact. Methods Five academic medical centers each using a different clinical chemistry analyzer system, collected their respective annual vendor purchase data (January 2022–December 2022). Consumables including reagent cartridges, packaging boxes, inserts associated with reagents, calibrators, and QC materials were categorized and weighed separately. Solid waste estimates for one million BMPs were calculated based on institutional testing volumes. Water and electricity usage for the analyzer systems was obtained and estimated CO 2 emissions (CO 2 e) were determined using standard emission factors and calculators. Results The study found that performing one million BMPs on wet chemistry analyzer platforms, Abbott, Beckman, Roche and Siemens generated 995 kg, 1,274 kg, 579 kg and 489 kg of solid waste respectively while the dry chemistry Ortho system produced 160,593 kg. The majority of waste across all platforms was plastic. Annualized CO 2 e from electricity and water use totaled to 69,665 kg CO 2 e. Conclusions Analysis of BMPs generates substantial amounts of solid waste and CO 2 e across major diagnostic platforms. These findings emphasize the urgent need for sustainability strategies in diagnostic laboratories to address their environmental footprint.

Objectives To compare the diagnostic performances of GFAP and UCH-L1 with S100B in detecting intracranial injury, while investigating the impact of confounding factors. Methods In a large prospective cohort of 1,010 patients with mild traumatic brain injury, we first evaluated the clinical performances of S100B and the GFAP/UCH-L1 combination. To explore the impact of pre-analytical interferences on GFAP and UCH-L1 levels, HIL indices (hemolysis, icterus, lipemia) were measured using the Atellica ® analyzer, and spiking experiments were performed with increasing concentrations of hemolysate, bilirubin, and Intralipid ® . We then assessed the influence of four confounders on biomarker specificity: age over 80 years, hemolysis, neurodegenerative diseases, and extracranial fractures. Finally, we evaluated the ability of the biomarkers to predict clinical outcomes at one month. Results S100B and the GFAP/UCH-L1 combination showed sensitivities of 96 and 100 %, and specificities of 25 and 27 %, respectively. False positives were significantly associated with age >80 and extracranial fractures for S100B; with age >80 and neurodegenerative diseases for GFAP; and with age >80, hemolysis, and extracranial fractures for UCH-L1. UCH-L1 levels were markedly increased by hemolysis, starting at 400 mg/L of hemoglobin. Age was the only confounding factor to significantly affect specificity. Using age-adjusted thresholds in patients over 80 increased specificity to 30 % for S100B and 33 % for GFAP/UCH-L1. Overall, the biomarkers exhibited limited predictive value and performed poorly for one-month clinical outcomes. Conclusions S100B and the GFAP/UCH-L1 combination demonstrated very high sensitivities, close to 100 %, with specificities of approximately 30 % for the diagnosis of intracranial lesions. Age-adjusted thresholds improve specificity in older patients, supporting their clinical implementation. This study also provides the first evidence that hemolysis significantly elevates UCH-L1 concentrations from 400 mg/L of hemoglobin.

Objectives Liquid chromatography-tandem mass spectrometry (LC–MS/MS) is increasingly used in clinical laboratories due to its high analytical specificity and multiplexing capabilities. While the pre-analytical and analytical phases have seen significant advances in automation, the post-analytical phase remains a bottleneck with limited standardisation. This survey aimed to identify gaps, benchmark practices against guidelines, and inform recommendations for improving consistency and quality in post-analytical data handling. Methods A descriptive electronic survey with 53 questions was distributed via the IFCC mailing list and collaborating organisations from May to July 2024. Questions covered participant characteristics, post-analytical quality indicators (e.g., chromatograms, calibration curves, ion ratios), quality assurance, and data management. Responses were benchmarked against three LC-MS/MS guidelines. Results Of 311 initial responses, 203 valid submissions from 57 countries were analysed. Laboratories reported diverse throughput: low (<100 samples/week, n=50), medium (100–2,000 samples/week, n=125), and high (>2,000 samples/week, n=28). Key findings included inconsistent acceptance criteria (e.g., variable ion ratios and signal-to-noise thresholds), reliance on manual data transcription (38 % of laboratories), and continued use of in-house spreadsheets. Practices often deviated from guidelines, with opportunities identified for harmonisation and automation. Conclusions In collaboration with the clinical mass spectrometry community, the IFCC-ETD developed six key recommendations from this survey to address challenges and enhance consistency, quality, and efficiency in post-analytical data handling.

Objectives Detection of paraneoplastic anti-neuronal antibodies (PNS autoantibodies) aids in diagnosing the particular syndrome and guides the search for an underlying tumor. To identify opportunities for harmonization that could enhance diagnostic value, we assessed variability in autoimmune serological testing for PNS across laboratories. Methods The European Autoimmunity Standardisation Initiative (EASI) developed a questionnaire addressing testing methods, digital tools, interpretation, clinical context, and quality assurance. This was distributed through two external quality assessment (EQA) providers (UK NEQAS and IfQ Lübeck) to clinical labs routinely performing PNS autoantibody assays. Results Among 139 responding laboratories, 78 % perform both cerebellum indirect immunofluorescence (IIF) and line/dot blot assays on serum and cerebrospinal fluid (76 %). Alignment on sample dilution (61 %) and conjugate type (70 %) is variable. Differences in antigen composition and reporting strategies contribute to variability in line/dot blot testing. Digitization is widespread for line/dot blot data (87 %) but limited for cerebellum IIF (31 %). About 53 % use testing algorithms that vary by sample type and requested antibodies. Internal quality control is performed by 89 % (IIF) and 48 % (line/dot blot), mainly using commercial controls. Nearly half (43 %) participate in multiple EQA programs; 46 % (IIF) and 42 % (line/dot blot) have ISO15189 certification, with 20 % planning certification. Positivity rates are low (<6 % in 73 % of labs), indicating low pre-test probability. While only 46 % restrict testing by discipline, most labs access clinical context (84 %) and interact with clinicians (63 %). Conclusions Considerable variability exists in PNS autoantibody assay execution, reporting, and quality control. Introducing lab-specific recommendations may harmonize practices and improve diagnostic quality.

Objectives To investigate the staining and serological characteristics of AC-30 pattern. Methods A total of 184 participants were recruited from patients who underwent routine antinuclear antibody testing between 2022 and 2023 at Peking Union Medical College Hospital. Cohort 1 (n=47) showed the AC-30 pattern on HEp-2 indirect immunofluorescence assay, and cohort 2 (n=137) showed AC-2 pattern as control. Anti-DFS70 antibody detection and DFS70 antigen adsorption assays were conducted. Pattern simulation assays were performed by combining serum samples exhibiting classic AC-2 pattern with other common HEp-2 IIFA patterns. Results Anti-DFS70 antibodies were detected in 97 % of patients in cohort one and in all patients in cohort 2. The titers of the IIFA pattern showed a weak correlation with anti-DFS70 antibody levels in cohort 1 (r=0.35, p=0.0331). In DFS70 antigen adsorption assays, a higher proportion of homogeneous nuclear staining was observed in cohort 1 (79 %) than in cohort 2 (62 %) (p=0.037). Simulated samples mixed classic AC-2 with homogeneous pattern resembled those of AC-30 pattern in both interphase and mitotic cells. Especially, the staining characteristics of AC-2 became increasingly indistinct when mixed with higher-titer homogeneous patterns. Among samples exhibiting homogeneous patterns post-DFS70 immunoadsorption, non-autoimmune conditions were more common in cohort one than cohort two. Conclusions The presence of relatively lower anti-DFS70 antibodies levels or coexisting high-titer homogeneous patterns may contribute to the development of the AC-30 pattern rather than AC-2. This finding needs to be further confirmed in larger-scale studies.

Objectives Neurofilament light (NfL) is an established biofluid marker of neuroaxonal injury for neurological diseases. Several high-throughput and sensitive immunoassays have been developed to quantify NfL in blood and cerebrospinal fluid (CSF), facilitating the use of NfL as a biomarker in research and clinical practice. However, because of the lack of rigorous comparisons of assays, it has been difficult to determine whether data are comparable and whether assay performance differs. Here, we compared the performance of five NfL immunoassays. Methods To assess the five NfL immunoassays (Fujirebio, ProteinSimple, Quanterix, Roche and Siemens), we used pooled plasma or pooled CSF, as well as unique samples from 20 healthy controls and 20 individuals with El Escorial defined probable or definite amyotrophic lateral sclerosis (ALS), to evaluate precision, parallelism and/or bias. We also examined correlations between plasma and CSF NfL concentrations within and across assays and evaluated their ability to differentiate healthy controls from individuals with ALS. Results Four of the five assays demonstrated exemplary performance based on our analyses of precision and parallelism. Across the five assays, NfL concentrations were lower in plasma than in CSF, although they displayed a high degree of correlation. We noted bias across assays; plasma NfL concentrations were lowest for the Roche assay and highest for the ProteinSimple assay. In addition, all assays reliably distinguished healthy controls from individuals with ALS using plasma or CSF NfL. Conclusions Four NfL assays demonstrated similar analytic performance. Alongside performance, other factors such as costs, accessibility, usability, footprint, and intended use, should be considered.

Objectives Automated urine particle analyzers are increasingly employed in clinical laboratories to standardize urinalysis. These systems can reduce workload, turnaround time and the inter-observer variation. This study aimed to assess the performance of five automated urine particle analyzers in detecting and categorizing dysmorphic erythrocytes (isomorphic-, dysmorphic erythrocytes, acanthocytes) and pathological casts, in particular erythrocyte casts, for screening of glomerular vs. non-glomerular hematuria, compared to manual phase-contrast microscopy, as the reference method. Methods The analytical performance of five automated urine particle analyzers, Atellica UAS 800, FUS-3000Plus, iQ200, SediMAX conTRUST Pro, and UF-4000, for measuring and categorizing erythrocytes and pathological casts was evaluated. Urine samples spiked with erythrocytes were used to assess precision and performance characteristics, while 68 urine samples from patients with glomerular hematuria were analyzed for concordance. Results Automated analyzers demonstrated acceptable precision at middle and high erythrocyte concentrations. At low levels, the iQ200 performed relatively well and the SediMAX met the manufacturer’s precision criteria. Manual on-screen review was required for 90 % of samples analyzed to achieve acceptable results for dysmorphic erythrocytes and cast detection. None of the analyzers exhibited strong agreement with manual microscopy for identifying isomorphic-, dysmorphic erythrocytes or acanthocytes (concordance <0.80), whereas the FUS-3000 and UF-4000 had relatively higher sensitivities for pathological cast detection (58.6 and 69.6 %, respectively). Conclusions Current automated urine particle analyzers are limited in accurately identifying dysmorphic erythrocytes and pathological casts, necessitating manual microscopy for reliable assessment. While automation offers potential for standardization and efficiency, significant technological advancements are required to improve diagnostic accuracy, reliability, and clinical applicability.

Objectives Identifying the potential sources of bias in the direct measurement of low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) is important. In this study, we aimed to investigate the effect of remnant cholesterol on LDL-C and HDL-C levels measured using homogenous methods. Methods We obtained 41 commutable frozen serum samples and measured LDL-C and HDL-C levels. Eight measurement systems were used, and the degree of bias was obtained by comparing with the values obtained using the reference methods. Correlations among remnant cholesterol/LDL-C, remnant cholesterol/HDL-C, and bias were analyzed using Spearman’s analysis. Results In all eight systems, samples with a positive bias >4 % had lower LDL-C levels and higher remnant cholesterol levels, as measured by the reference methods, compared to those with a bias ≤4 %. A significant correlation between remnant cholesterol/LDL-C and a positive bias of LDL-C was observed in six of the eight systems evaluated. For HDL-C bias, three systems showed a positive correlation, and three systems showed a negative correlation. In some systems, LDL-C bias was higher in samples with remnant cholesterol/LDL-C ≥0.25 than in those with remnant cholesterol/LDL-C <0.25. Conclusions Remnant cholesterol has a potential effect on direct LDL-C and HDL-C measurements, which has been observed when several measurement systems are used. For these systems, manufacturers should improve the methods to reduce the interference of remnant cholesterol.

Objectives Dental amalgam (50 % mercury (Hg) by weight) is a commonly used material to restore a tooth damaged by decay. In recent years, amalgam safety has become a matter of ongoing controversy. This hypothesis-testing epidemiological study evaluated the relationship between blood Hg concentrations and amalgams in American adults. Methods Examination of the 2015–2016 National Health and Nutrition Examination Survey (NHANES) was undertaken using SAS, version 9.4 (Cary, NC, USA), survey regression statistical modeling (with adjustments for covariates). A total of 180,811,187 weighted-Americans (n=1,377) between the ages of 18–70 years-old, with known: dental filing surface status; urinary Hg concentrations; total and blood Hg species (inorganic and methyl-Hg) concentrations; bodyweight; and urine flow rates were examined. Results Significant increases were found in the blood concentrations of total and inorganic Hg, when comparing adults exposed to amalgams as compared to adults not exposed to amalgams. Amalgam surfaces significantly correlated with blood inorganic Hg concentrations, and estimated daily Hg vapor doses from amalgams significantly correlated with blood total and inorganic Hg concentrations. This study supports the importance of blood as an important transport avenue for Hg, which is dose-dependently released by amalgams, to accumulate in tissues and cells throughout the body. Conclusions Persons with amalgams, desiring to lower their blood Hg concentrations, should consult with a dentist trained and certified in safe amalgam removal. Also, pharmaceutical treatments to reduce/render non-toxic the blood Hg concentrations from amalgams should be considered. Efforts should be made to reduce/eliminate the continued use of amalgams.

Objectives High-sensitivity cardiac troponin T (hs-cTnT) is used in pediatric healthcare to diagnose and assess the prognosis of various cardiac diseases, such as acute myocarditis, congenital heart disease, dilated cardiomyopathy, and heart failure. However, the standardized age-related reference intervals, 99th percentile cut-offs, and clinical guidelines are currently unavailable, especially in newborns and infants, making interpreting this biomarker challenging. Methods Residual serum samples were collected from 1,047 pediatric subjects with no cardiovascular disease, including 145 neonates and 902 infants. The concentrations of hs-cTnT were analyzed using Roche’s Elecsys Troponin T hs assay. The age-related 99th percentile cut-offs with 90 % confidence intervals (CIs) of hs-cTnT were established. Results Subjects were divided into age subgroups as follows: newborns (0–30 days, n=142), infants aged 1 month (31–60 days, n=223), 2 months (61–90 days, n=152), 3–4 months (91–150 days, n=211), 5–6 months (151–210 days, n=149), and 7 months-1 year (211–365 days, n=149). Serum hs-cTnT concentrations were highest during the first month of life and progressively declined in a year. The 99th percentile cut-offs of hs-cTnT concentration in each group were as follows: 114 (90 % CI: 114-114), 65.6 (63.7–66.1), 55.2 (54.4–55.2), 30.4 (29.1–30.9), 23.5 (22.1–23.5), 12.7 (12.4–12.7) ng/L. The 97.5th percentile cut-offs for each group were 109.4 (105.2–114), 62.5 (60.5–63.9), 52.3 (50.3–54.4), 29 (28.5–29.1), 20.5 (19.2–23.4), and 12 (12–12.1) ng/L. Conclusions This study aimed to provide reliable pediatric reference values for hs-cTnT based on a population of well-child children. These reference intervals and 99th percentile cut-offs will inform clinical decisions in the pediatric cardiology setting.

Objectives Neuron-specific enolase (NSE) is a clinically relevant biomarker used in the assessment of neuronal damage and in the diagnosis and monitoring of certain cancers. Despite its diagnostic importance, paediatric-specific reference intervals (RIs) for NSE are currently lacking. This study aimed to establish paediatric RIs for NSE in serum and to evaluate the influence of preanalytical factors on NSE measurements. Methods Residual serum samples from routine allergy testing in 242 Danish children (aged 0.1–17.9 years) were analysed using the Roche Elecsys ® NSE assay on a Cobas platform. Both traditional non-parametric, age-partitioned RIs and continuous RIs derived via quantile regression were established. In addition, we assessed the impact of preanalytical variables, including haemolysis, and evaluated a previously proposed correction method for haemolysed samples within this cohort. Results Both the non-parametric and continuous approaches yielded consistent RIs, showing an age-dependent decline in serum NSE concentrations irrespective of sex. The traditional age-partitioned RI (95th percentile, one-sided) indicated upper limits of 36.9 μg/L and 32.0 μg/L for the age groups 0–5 and 6–17 years of age, based on samples with haemolysis <10 mg/dL haemoglobin. Conclusions This study defines age-specific paediatric RIs for serum NSE, demonstrating a physiological decline with age and highlighting higher NSE levels in healthy children compared to adults. Furthermore, within a limited range, a previously proposed simple linear correction method was validated for adjusting NSE values in mildly haemolysed samples using the newly established RIs.

Objectives To analyze the performance of laboratories in the China National External Quality Assessment Scheme (China NEQAS) for HbA 2 and HbF testing, and to propose strategies for quality improvement. Methods Data were obtained from China NEQAS from 2013 to 2024 using both commercially lyophilized controls and in-house whole blood controls. The distributions of methods and instruments were analyzed. Inter-laboratory coefficient variations (CVs) and target values were compared between two types of controls and between method-instrument platforms over 12 years. Results The in-house controls remained homogeneous and stable for almost one month at 2–8 °C and for one year at −80 °C. Capillary electrophoresis (CE) became the dominant method, adopted by 84.3 % of labs in 2024. For HbA 2 , two EQA controls had comparable concentration ranges and inter-laboratory CVs. HbF in-house controls covered broader pathological concentrations than commercial ones. CE demonstrated lower inter-laboratory CVs for both analytes: HbA 2 was 2.1 ± 1.8 % vs. 5.5 ± 4.8 % (high-performance liquid chromatography, HPLC) and 6.5 ± 4.1 % (agarose gel electrophoresis, AGE), while HbF was 3.2 ± 1.9 % vs. 5.0 ± 1.6 % (HPLC) and 8.6 ± 6.8 % (AGE). Significant discrepancies in maximum-to-minimum mean concentrations were observed among different method-instrument platforms when testing the same controls (up to 10 % for HbF). Conclusions In-house controls demonstrate homogeneity, stability and intrinsic commutability, and have an expanded concentration range, can serve as a reliable alternative to commercial controls for EQA. High-precision techniques such as CE should be favoured. Furthermore, the development of reference methods and commutable reference materials is essential for standardization.

Objectives The European Organization for External Quality Assurance Providers in Laboratory Medicine (EQALM) Haematology Working Group (WG) has previously provided two guidelines to support the standardization of hematology blood smear external quality assessment (EQA) (Vives CJL, Albarède S, Flandrin G, Heller S, Horvath K, Houwen B, et al. Guidelines for blood smear preparation and staining procedure for setting up an external quality assessment scheme for blood smear interpretation. Part I: control material. Clin Chem Lab Med 2004;42:922–6; Vives CJL, Van Blerk M, Albarède S, Gutierrez G, Heller S, Nordin G, et al. Guidelines for setting up an external quality assessment scheme for blood smear interpretation. Part II: survey preparation, statistical evaluation and reporting. Clin Chem Lab Med 2006;44:1039–43) but these recommendations did not include analytical performance specifications. In this paper the WG provides advice on the performance specifications for the evaluation of blood cell identification, comments on morphological characteristics and diagnostic hypothesis. Methods To develop these specifications, the WG made a survey of the practices in use by EQALM members and provided a standard set of EQA data, provided by one of the members, for performance evaluation. Results The results of this exercise show a variation in the performance assessment outcomes from one EQA provider to another, suggesting that a degree of standardization would be of benefit to participating laboratories. Conclusions The WG has provided advice on common performance specifications, based on model 1 of the Milan Consensus (Sandberg S, Fraser CG, Horvath AR, Jansen R, Jones G, Oosterhuis W, et al. Defining analytical performance specifications: consensus statement from the 1st strategic conference of the European federation of clinical chemistry and laboratory medicine. Clin Chem Lab Med 2015;53:833–5).

Objectives Ocular lymphomas are rare diseases that occur in ocular adnexa or specific intraocular regions. Although histology remains the gold standard, cytology and flow cytometry (FCM) are fundamental, mainly when a biopsy sampling is technically demanding. This study evaluated the effectiveness of FCM in diagnosing ocular diseases, especially hematologic malignancies, and compared its performance with the other techniques. Methods In this monocentric study we collected patient characteristics from 165 patients referred to Padua University Hospital. Sensitivity, specificity, positive (PPV) and negative predictive values (NPV), accuracy and concordance rate were evaluated for cytologic, FCM, and/or histologic analyses performed. Results Patients were classified as having hematologic neoplasms (n=86), non-hematologic neoplasms (n=33) or non-neoplastic diseases (n=46). Hematologic neoplasms comprised 74 cases with ocular adnexal hematologic lesions and 12 cases with intraocular lymphomas and leukemias. FCM analysis of ocular specimens allowed valuable results in 71/86 hematologic cases, failure being mainly due to absence of cellularity (8/15). Histology showed 100 % PPV for both hematologic and non-hematologic neoplasms; FCM reached 97 and 66 %, and cytology 80 and 91 %, respectively. Concordance between FCM and cytology was lower (κ<0.61) than with histology (κ=0.677). FCM phenotype matched ocular lymphoma histotype in 95.8 % of cases. The lymphocytic infiltrate by FCM was 39.5 % in non-neoplastic, 64 % in non-hematologic and 89.5 % in hematologic diseases with B lymphocyte prevalence of 31.3 %, 21.0 % and 93.3 %, respectively. Clonality analysis remains essential regardless of these values. Conclusions FCM provides rapid results, studies numerous markers simultaneously, and analyzes ocular aspirates from minimally invasive samples. For this reason, it is desirable to permanently integrate this technique into the diagnostic process of ocular disorders, particularly when hematologic malignancies are suspected, as part of an integrated multidisciplinary pathway.

Objectives The potential relationship between cardiac troponin (cTn) composition and etiologies of myocardial injury may provide diagnostic insights for myocardial infarction (MI). We compared the performance of novel ternary cTnI–cTnT–TnC complex (ITC) assays with high-sensitivity (hs)-cTnI and hs-cTnT in differentiating MI patients at presentation. Methods Plasma samples from 1,210 patients with suspected MI were collected. Major cTn forms were quantified using long-cTnT ITC complex, hs-total ITC complex, hs-cTnI and hs-cTnT assay. We compared the assay performance based on the 99th percentile upper reference limits (URLs), evaluated their area under the receiver operating characteristic curve (AUC), and assessed their ability to differentiate MI using established cut-off values. Results A total of 138 (11.4 %) patients were diagnosed with MI. 86 % patients had long-cTnT ITC complex concentration below the 99th percentile URLs (sensitivity: 0.818, specificity: 0.943, NPV: 0.975, PPV: 0.649). AUC was 0.959 for long-cTnT ITC complex, comparable to hs-cTnI (0.953) and superior to hs-total ITC complex (0.889) and hs-cTnT (0.927). Using established cut-off values, the long-cTnT ITC complex assay classified 10 % of patients as rule-in and 52 % as rule-out (sensitivity: 0.993, NPV: 0.998, specificity: 0.972, PPV: 0.760), which was comparable to hs-cTnI and better than hs-total ITC complex and hs-cTnT. In early-presenting patients, the long-cTnT ITC complex demonstrated enhanced discrimination and improved diagnostic performance. Conclusions The long-cTnT ITC complex assay demonstrated performance comparable to hs-cTnI and superior to hs-total ITC complex and hs-cTnT in distinguishing MI using a single sample at presentation. These findings suggest its potential utility in improving risk stratification and triage strategies, particularly among early-presenting patients.