BACKGROUND:Vitamin D is a key regulator of skeletal homeostasis, and hypovitaminosis D is highly prevalent among postmenopausal women, who are at increased risk of osteoporosis, sarcopenia, and related complications. Accurate assessment of serum 25-hydroxyvitamin D [25(OH)D] is therefore essential. However, substantial variability exists among analytical methods, particularly between automated chemiluminescent immunoassays (CLIA) and liquid chromatography-tandem mass spectrometry (LC-MS/MS), the latter considered the reference technique. This study aimed to compare four analytical methods, three CLIA platforms, and LC-MS/MS for measuring circulating 25(OH)D levels in a cohort of menopausal and postmenopausal women. METHODS:A total of 425 serum samples from menopausal and postmenopausal women representing the real-world distribution of vitamin D levels in this population were analyzed using three automated CLIA systems and LC-MS/MS. Method comparison, agreement, precision through quality control assessment, total error, and sigma were evaluated. RESULTS:The evaluated CLIA platforms (Abbott, Snibe, and Siemens) showed strong correlation with LC-MS/MS, with r = 0.919, r = 0.978, and r = 0.879. Furthermore, all assays showed excellent precision (CV < 5%), with good-to-acceptable total error (TE) and Sigma-metric performance. CONCLUSIONS:In conclusion, these findings demonstrate that while CLIA platforms offer a reliable and precise alternative for routine clinical use, these findings underscore the importance of method selection and result interpretation in the clinical assessment of vitamin D status in postmenopausal women. Furthermore, it highlights the ongoing need to minimize inter-assay variability and ensure consistent vitamin D assessment.

Objectives Faecal calprotectin levels closely reflect the quantity of neutrophils within the intestinal lumen, enabling the sensitive detection of acute inflammatory responses in the gut. A novel, automated chemiluminescent immunoassay (CLIA) was developed for the quantification of faecal calprotectin.Methods All CHORUS CLIA Calprotectin reagents, both reagents and the recombinant protein used for the calibration of the test, were produced in-house. To validate the assay, the limit of detection (LoD) and quantification (LoQ), precision, and reliability were determined. The assay was compared with the Calprest (R) NG (Eurospital n. 9,069) by Spearman's correlation coefficient, Passing-Bablok, and Bland-Altman analyses.Results The assay was linear in the concentration range between the LoQ, and no differences emerged in the intra-assay and inter-assay precision; the LoD was 22.2 mg/kg. Compared to Calprest (R), CHORUS CLIA Calprotectin showed excellent agreement: the Passing Bablok equation was y=-0.5177 + 0.9954x with a Spearman correlation coefficient of 0.98 (95 %CI: 0.97-0.98). In the Bland-Altman test, the mean difference was -4.38 (95 %CI: -18.174-9.408) with upper and lower limits of 135.96 and -144.73, respectively.Conclusions CHORUS CLIA Calprotectin was accurate and reliable in determining the concentration of faecal calprotectin and may become a useful tool in clinical practice. The assay proves to be highly precise, robust, and capable of accurately measuring calprotectin across the full clinical range, reliably distinguishing between physiological and pathological levels. Comparative analysis confirms that the CHORUS CLIA Calprotectin assay delivers results in strong agreement with Calprest NG used as the reference method.

Laser-induced graphene (LIG) is a novel, low-cost material with excellent electrical properties that has recently gained increasing attention in bioengineering for both sensing and actuation applications. However, its integration into light microscopy-compatible platforms for in vitro biological studies, such as lab-on-chip systems, is often hindered by complex and potentially invasive techniques for transferring it into final substrates. In this work, we propose a novel approach for the direct fabrication of LIG on polystyrene substrates using commercial polyimide adhesive tape, CO 2 laser irradiation, and a simple peel-off process, enabling the production of fully in vitro-compatible devices. The material is comprehensively characterized through scanning electron microscopy (SEM), Raman spectroscopy, electrical resistivity measurements, finite element method (FEM) simulations, and machine learning based analysis. The resulting LIG electrodes are integrated into a muscle-on-chip microfluidic device, where they successfully generated electrical stimuli, inducing contractions in differentiated myotubes. These contractions are monitored by time-lapse microscopy and quantitatively assessed using video analysis, demonstrating the tissue response in phase with electrical stimulation.

Abstract Introduction Herpes simplex virus type 1 (HSV-1) is highly prevalent worldwide, making accurate serological testing essential for both clinical diagnosis and epidemiological surveillance. Automated chemiluminescent immunoassays (CLIAs) offer operational advantages over enzyme-linked immunosorbent assays (ELISAs); however, their diagnostic performance relative to Western blot (WB) confirmation in high-prevalence settings remains insufficiently characterized. Hypothesis/Gap Statement The comparative diagnostic accuracy of CLIA- and ELISA-based assays for HSV-1 IgG detection, when benchmarked against a WB reference standard in endemic populations, remains unclear. Aim This study aimed to evaluate HSV-1 IgG seroprevalence and diagnostic performance of one CLIA and two ELISA platforms using Western blot as the reference method. Methodology Four hundred archived serum samples from adult male craft and manual workers in Qatar were tested using the Mindray CL-900i CLIA, HerpeSelect ELISA, NovaLisa ELISA, and Euroimmun Western blot. Seroprevalence, diagnostic accuracy, and interassay agreement were assessed using WB as the reference standard, with equivocal and indeterminate results excluded from analysis. Results HSV-1 IgG seroprevalence estimates were comparable across assays: HerpeSelect 72.5%, Mindray 70.5%, NovaLisa 66.3%, and Western blot 66.5%, with no statistically significant differences (all p > 0.05). The Mindray CLIA demonstrated the highest diagnostic performance (sensitivity 95.7%, specificity 88.9%, accuracy 93.4%) and strong agreement with Western blot (κ = 0.85). HerpeSelect showed substantial agreement (κ = 0.81), while NovaLisa exhibited lower specificity. Conclusion CLIA- and ELISA-based assays produced comparable HSV-1 seroprevalence estimates in this high-prevalence population; however, diagnostic accuracy varied across platforms. The CLIA platform demonstrated the strongest agreement with Western blot, supporting its use in high-throughput settings, while confirmatory testing remains important to minimize misclassification. Key Points What is known: HSV-1 serological diagnosis relies mainly on ELISA assays, while automated CLIA platforms are increasingly used in high-throughput laboratories but remain insufficiently evaluated against Western blot confirmation. What is new: This study provides a large head-to-head comparison of CLIA and ELISA platforms for HSV-1 IgG detection using Western blot as the reference standard in a high-prevalence population. Clinical implications: Automated CLIA systems demonstrated strong diagnostic accuracy and may represent reliable high-throughput alternatives for HSV-1 serological screening in clinical laboratories. Impact Statement Accurate serological diagnosis of herpes simplex virus type 1 (HSV-1) is essential for clinical management, epidemiological surveillance, and public health decision-making, particularly in populations where infection is highly prevalent. This study adds to the existing literature by providing a large, head-to-head comparison of automated chemiluminescent immunoassay (CLIA) and enzyme-linked immunosorbent assay (ELISA) platforms for HSV-1 IgG detection, benchmarked against Western blot confirmation in a real-world, high-prevalence setting. By demonstrating that different serological platforms can yield similar population-level seroprevalence estimates yet differ in diagnostic accuracy and specificity, this work highlights the risk of misclassification when confirmatory testing is not considered. The findings are of broad relevance to clinical microbiology laboratories, diagnostic services, and public health surveillance programs that rely on serological assays for HSV-1 screening. The study represents an incremental but important step in refining assay selection and interpretation, supporting more reliable laboratory diagnostics and improved understanding of HSV-1 infection burden in endemic populations. Data Availability Statement The data that support the findings of this study are available from the corresponding author upon reasonable request.

Background: Metabolic syndrome (MetS) patients have impaired hypothalamic regulatory functions involved in food intake and energy expenditure and suffer from a state of meta-inflammation. Pre-clinical studies demonstrated that ultramicronized palmitoylethanolamide (PEA) acts both on the adipose tissue and the central nervous system, while hydroxytyrosol (HTyr) counteracts several types of dysmetabolism. Objectives: The aim of our randomized crossover double-blind placebo-controlled pilot study was to evaluate the potential effects of a food supplement (FS) containing a co-micronized formulation of PEA and rutin along with HTyr, combined with a tailored calorie-controlled Mediterranean diet, in patients with MetS. Methods: Nineteen patients were enrolled and block-randomized to an eight-week MD together with the FS or placebo. After a two-week washout period, the treatments were reversed. Data on laboratory parameters and those detected by capillary sampling, anthropometry, body composition analysis, ultrasound examination, blood pressure monitoring, the 36-Item Short-Form Health Survey questionnaire, handgrip strength test, and physical performance tests were collected at each time point (protocol code R.S. 262.22, registered on 20 December 2022). Results: At the end of the study, patients supplemented with the FS showed a significant reduction in body weight, body mass index, fat mass, and inflammation biomarkers (CRP and ESR), compared to placebo-supplemented patients. In contrast, the fat-free mass, phase angle, and body cell mass were increased in FS compared to placebo patients. Conclusions: Although preliminary, the results of our clinical study suggest that co-micronized PEA–rutin and HTyr may be of help against adiposopathy in patients with MetS.

Motor complications (MC), including fluctuations, represent a disabling milestone of Parkinson’s disease (PD) course, although the underlying early pathophysiological mechanisms remain unclear. We therefore investigated whether the biological profile at PD onset, as defined through a panel of CSF biomarkers, may predispose to the development of MC. We conducted a dual-center retrospective longitudinal study involving 131 de novo (DN) PD patients (newly diagnosed, untreated). At baseline, patients were evaluated by motor and non-motor scores, and the measurement of CSF total α-synuclein (α-syn), total and phosphorylated-181-tau (t-tau, p-tau), amyloid-β42 and amyloid-β40 (Aβ42, Aβ40) levels, p-tau/t-tau, Aβ42/Aβ40, and p-tau/Aβ42 ratios. According to the successive development of MC, patients were classified as “with MC” (wMC) or “without MC” (noMC). A control group of 107 controls was also collected. Variables were compared between groups, adjusting for main covariates; ROC and Cox analyses evaluated predictive values. The DN PD cohort was followed for 57 (± 18) months, with 38 (29

The increasing global burden of chronic kidney disease (CKD) necessitates the development of simple and inexpensive diagnostic tools. Capitalizing on the relationship between breath composition and CKD, we introduce a disposable array of four resistive gas sensors printed on a low-cost, disposable substrate and embedded in the internal layers of FFP2 facemasks. Sensors are based on blends of porphyrins─a molecular family often used in breath analysis─and the PEDOT/PSS conducting polymer. The individual sensors demonstrate remarkable sensitivity to ammonia and other CKD-related volatile compounds, while the combinatorial selectivity of the sensor array enables the identification of volatile compounds regardless of their concentration. The diagnostic capabilities of the device were tested on a cohort of CKD patients and a control group. To address the absence of a reference gas inside the facemask, we developed a measurement protocol based on breathing cycles at different rates. The application of a continuous wavelet transform to the sensor signals produces stable and reproducible features. Linear Discriminant Analysis of sensor features achieved the identification of CKD patients with 93.3% true positives and 86.7% true negatives. Additional evidence suggests that the sensor array can stratify CKD patients according to the severity of renal dysfunction, indicating its potential use in monitoring disease progression.

Objective Serum amyloid A (SAA) is a protein involved in the acute phase of inflammation. SAA expression is upregulated in humans during the acute phase of various viral infections; in addition, SAA can be a useful biomarker to predict the severity and prognosis of COVID-19 patients. This study aimed to evaluate a new chemiluminescence test for SAA detection. Methods All serum samples were measured for SAA on a Maglumi 800 (Snibe, Shenzhen, China) and compared with a BN ProSpec (Siemens, Munich, Germany) in the routine of the clinical laboratory of the University Hospital of the Tor Vergata University of Rome (Rome, Italy). Analytical precision, the correlation coefficient, and linearity were assessed. Statistical analyses were performed. Results The linearity test was performed via serial dilutions and revealed a correlation coefficient equivalent to 0.9998. The results of the Snibe SAA test correlated well with those obtained by the SAA Siemens test, with a correlation coefficient of 0.974 (P ​< ​0.001). The intra- and interrun precision, as well as carryover, were assessed. Conclusions The results obtained from this study demonstrated that the new Snibe SAA test has reliable analytical performance and good accuracy and could represent a valid tool for routine hospital laboratory analysis.

Kidney transplantation is currently the treatment of choice for patients with end-stage kidney diseases. Although significant advancements in kidney transplantation have been achieved over the past decades, the host’s immune response remains the primary challenge, often leading to potential graft rejection. Effective management of the immune response is essential to ensure the long-term success of kidney transplantation. To address this issue, immunosuppressives have been developed and are now fully integrated into the clinical management of transplant recipients. However, the considerable inter- and intra-patient variability in pharmacokinetics (PK) and pharmacodynamics (PD) of these drugs represents the primary cause of graft rejection. This variability is primarily attributed to the polymorphic nature (genetic heterogeneity) of genes encoding xenobiotic-metabolizing enzymes, transport proteins, and, in some cases, drug targets. These genetic differences can influence drug metabolism and distribution, leading to either toxicity or reduced efficacy. The main objective of the present review is to report an historical overview of the pharmacogenetics of immunosuppressants, shedding light on the most recent findings and also suggesting how relevant is the research and investment in developing validated NGS-based commercial panels for pharmacogenetic profiling in kidney transplant recipients. These advancements will enable the implementation of precision medicine, optimizing immunosuppressive therapies to improve graft survival and kidney transplanted patient outcomes.

Background: Urine microbial analysis is a frequently requested test that is often associated with contamination during specimen collection or storage, which leads to false-positive diagnoses and delayed reporting. In the era of digitalization, machine learning (ML) can serve as a valuable tool to support clinical decision-making. Methods: This study investigates the application of a simple artificial neural network (ANN) to pre-identify negative and contaminated (false-positive) specimens. An ML model was developed using 8181 urine samples, including cytology, dipstick tests, and culture results. The dataset was randomly split 2:1 for training and testing a multilayer perceptron (MLP). Input variables with a normalized importance below 0.2 were excluded. Results: The final model used only microbial and either urine color or urobilinogen pigment analysis as inputs; other physical, chemical, and cellular parameters were omitted. The frequency of positive and negative specimens for bacteria was 6.9% and 89.6%, respectively. Contaminated specimens represented 3.5% of cases and were predominantly misclassified as negative by the MLP. Thus, the negative predictive value (NPV) was 96.5% and the positive predictive value (PPV) was 87.2%, leading to 0.82% of the cultures being unnecessary microbial cultures (UMC). Conclusions: These results suggest that the MLP is reliable for screening out negative specimens but less effective at identifying positive ones. In conclusion, ANN models can effectively support the screening of negative urine samples, detect clinically significant bacteriuria, and potentially reduce unnecessary cultures. Incorporating morphological information data could further improve the accuracy of our model and minimize false negatives.

Emerging evidence highlights a possible interplay between serum lipid profiles and Parkinson’s disease (PD), but the biological underpinnings remain largely unexplored. In this cross-sectional study, we investigated whether serum lipid levels (total cholesterol, high-density lipoprotein cholesterol (HDL), non-high-density lipoprotein cholesterol, and triglycerides) were associated with clinical severity and cerebrospinal fluid (CSF) biomarkers in early-stage PD patients. A cohort of 90 PD patients and 74 matched controls underwent serum lipid and CSF biomarker assessment and APOE genotyping. While serum lipid levels did not differ significantly between groups, PD patients showed reduced CSF α-synuclein. Notably, higher HDL levels correlated with higher CSF α-synuclein and amyloid-β42 (Aβ42) concentrations and milder motor impairment, independent of APOE ε4 status. APOE ε4 carriers displayed increased CSF phosphorylated tau and reduced Aβ42/Aβ40 ratio, but APOE genotype did not modify the observed HDL associations. These findings suggest that higher circulating HDL levels are associated with a milder clinical phenotype and a more favorable CSF biomarker profile in early-stage PD, potentially reflecting a protective role independent of APOE genotype. Further studies are warranted to validate these observations and to assess their therapeutic implications.

Depending on age, both the risk and characteristics of Parkinson's disease (PD) differ between the sexes. The immune system might have a role; however, human-based evidence remains scarce. Here, we investigated the relationship between peripheral immune cellular composition and the clinical-biological sexual dimorphism of PD. The leukocyte population count (neutrophils, lymphocytes, monocytes, eosinophils, and basophils), the neutrophil-to-lymphocyte ratio (NLR), and the monocytes-to-lymphocytes ratio (MLR) were collected and compared in 117 PD patients and 86 controls (CTLs), and then related to blood levels of sex hormones, CSF markers of neurodegeneration (α-synuclein, amyloid-β-42, amyloid-β-40, total tau, and phosphorylated-181-tau), and clinical features in male and female PD patients. Finally, a cluster analysis based on the three main leukocyte populations (neutrophils, lymphocytes, monocytes) was performed for the entire PD cohort. Male PD patients had lower lymphocyte counts and higher NLR than male CTLs. Females with PD had lower monocyte counts, NLR, and MLR than males with PD. Lymphocyte counts correlated with cognition in male, but not female, PD patients. Finally, two clusters of peripheral immune cellular composition were identified: the "high peripheral inflammation" one, mostly comprising male patients, with worse clinical features and greater central α-synuclein burden, and the "low peripheral inflammation cluster", which mainly comprised female patients, with milder clinical features and lower central synucleinopathy. In conclusion, the peripheral immune pattern entails sex-specific clinical-biological profiles in PD. Moreover, systemic inflammation clusters with sex, sexual hormones, clinical features, and central synucleinopathy in PD, supporting the relevance of immunity in sexual dimorphism of the disease.

Background: 25-Hydroxy vitamin D [25(OH)D] is the circulating form of vitamin D. Its deficiency is a major global health concern, affecting over one billion people. Beyond its role in bone health, low vitamin D levels have been implicated in a wide range of chronic and inflammatory diseases, including diabetes, chronic kidney disease, and cancer. While immunoassays are widely used in routine testing, liquid chromatography–tandem mass spectrometry (LC-MS/MS) remains the reference method for its superior accuracy. This study aimed to evaluate the analytical performance of the Autobio 25(OH)D chemiluminescence assay (Autobio Diagnostics, Zhengzhou, China) compared with LC-MS/MS (Chromsystems Instruments & Chemicals GmbH, Gräfelfing, Germany) and the Siemens chemiluminescent microparticle immunoassay (Siemens HealthCare, Erlangen, Germany). Additionally, the influence of age and sex on 25(OH)D concentrations was examined to explore potential demographic and pathophysiological variations. Methods: 200 residual serum samples were analyzed to compare all three methods. Precision and linearity were verified. Statistical analysis was performed. Results: The Autobio assay showed good correlation with LC-MS/MS (R2 = 0.953; p < 0.001), with acceptable bias and precision (CV < 10%) and confirmed linearity. Age- and sex-related differences were observed, indicating demographic influences on vitamin D status. Conclusions: Accurate and accessible laboratory testing for 25(OH)D is therefore essential for both disease prevention and clinical management. The Autobio 25(OH)D assay demonstrated strong correlation with LC-MS/MS and high analytical reliability. Its good performance makes it a valuable tool for routine assessment of 25(OH)D and for supporting the early detection or monitoring of hypovitaminosis D in clinical practice.

Background: International guidelines recommend the use of high-sensitivity cardiac troponin (hs-cTn) I and T methods for the detection of myocardial injury as a pre-requisite for the diagnosis of acute myocardial infarction (AMI) in patients admitted to the emergency department. Recently, Mindray (Mindray Bio-Medical Electronics Co., Ltd., Shenzhen, China) has introduced a new chemiluminescence immunoassay (CLIA) for the detection of the cTn complex. The present study aims to verify and validate the hs-cTnI Mindray assay on the new automated CL2600i analyzer compared to the routine Alinity-i series instrument by Abbott (Abbott, Chicago, IL, USA). Methods: This study evaluated linearity, precision through the 5 × 5 protocol, methodological comparison on plasma and serum matrices, hs-cTnI 99th percentile imprecision, and the hs-cTnI detection rate in a healthy population. Results: The results obtained proved that the performance of the Mindray hs-cTnI test on the CL2600i platform was closely comparable to the Abbott Alinity-i system (plasma R2: 0.974; serum R2: 0.995). The CVs were consistently low, and no significant differences were reported. Excellent analytical performance, with high sensitivity, was also observed in the healthy population (overall detection rate: 79%), as well as good linearity within the measuring range (R2: 0.994). Conclusions: The Mindray hs-cTnI test confirms its robustness and utility in routine practice as an advanced assay. The new technology, with more sensitive detection methods, may improve the accuracy and reliability of cardiac biomarker testing, ultimately leading to better outcomes in the management of patients with AMI and other cardiac conditions.

Early-onset Parkinson’s disease (EOPD) occurs during the fertile life, when circulating neuroactive sex hormones might enhance the sexual dimorphism of the disease. Here, we aimed to examine how sex hormones can contribute to sex differences in EOPD patients. A cohort of 34 EOPD patients, 20 males and 14 females, underwent comprehensive clinical evaluation of motor and non-motor disturbances. Blood levels of estradiol, total testosterone, follicle-stimulating hormone, and luteinizing hormone were measured in all patients and correlated to clinical features. We found that female patients exhibited greater non-motor symptoms and a relatively higher rate of dystonia than males. In females, lower estradiol levels accounted for higher MDS-UPDRS-II and III scores and more frequent motor complications, while lower testosterone levels were associated with a major occurrence of dystonia. In male patients, no significant correlations emerged. In conclusion, this study highlighted the relevance of sex hormone levels in the sexual dimorphism and unique phenotype of EOPD.

ABSTRACTBackgroundPriming with ChAdOx1 followed by heterologous boosting is considered in several countries. Nevertheless, analyses comparing the immunogenicity of heterologous booster to homologous primary vaccination regimens and natural infection are lacking. In this study, we aimed to conduct a comparative assessment of the immunogenicity between homologous primary vaccination regimens and heterologous prime‐boost vaccination using BNT162b2 or mRNA‐1273.MethodsWe matched vaccinated naïve (VN) individuals (n = 673) with partial vaccination (n = 64), primary vaccination (n = 590), and primary series plus mRNA vaccine heterologous booster (n = 19) with unvaccinated naturally infected (NI) individuals with a documented primary SARS‐CoV‐2 infection (n = 206). We measured the levels of neutralizing total antibodies (NTAbs), total antibodies (TAbs), anti‐S‐RBD IgG, and anti‐S1 IgA titers.ResultsHomologous primary vaccination with ChAdOx1 not only showed less potent NTAb, TAb, anti‐S‐RBD IgG, and anti‐S1 IgA immune responses compared to primary BNT162b2 or mRNA‐1273 vaccination regimens (p < 0.05) but also showed ~3‐fold less anti‐S1 IgA response compared to infection‐induced immunity (p < 0.001). Nevertheless, a heterologous booster led to an increase of ~12 times in the immune response when compared to two consecutive homologous ChAdOx1 immunizations. Furthermore, correlation analyses revealed that both anti‐S‐RBD IgG and anti‐S1 IgA significantly contributed to virus neutralization among NI individuals, particularly in symptomatic and pauci‐symptomatic individuals, whereas among VN individuals, anti‐S‐RBD IgG was the main contributor to virus neutralization.ConclusionThe results emphasize the potential benefit of using heterologous mRNA boosters to increase antibody levels and neutralizing capacity particularly in patients who received primary vaccination with ChAdOx1.