Search Results (64)

Background/Objectives: The clinically validated multiplex Oncuria bladder cancer (BC) assay quickly and noninvasively identifies disease risk and tracks treatment success by simultaneously profiling 10 protein biomarkers in voided urine samples. Oncuria uses paramagnetic bead-based fluorescence multiplex technology (xMAP^®; Luminex, Austin, TX, USA) to simultaneously measure 10 protein analytes in urine [angiogenin, apolipoprotein E, carbonic anhydrase IX (CA9), interleukin-8, matrix metalloproteinase-9 and -10, alpha-1 anti-trypsin, plasminogen activator inhibitor-1, syndecan-1, and vascular endothelial growth factor]. Methods: In a pilot study (N = 36 subjects; 18 with BC), Oncuria performed essentially identically across three different common analyzers (the laser/flow-based FlexMap 3D and 200 systems, and the LED/image-based MagPix system; Luminex). The current study compared Oncuria performance across instrumentation platforms using a larger study population (N = 181 subjects; 51 with BC). Results: All three analyzers assessed all 10 analytes in identical samples with excellent concordance. The percent coefficient of variation (%CV) in protein concentrations across systems was ≤2.3% for 9/10 analytes, with only CA9 having %CVs > 2.3%. In pairwise correlation plot comparisons between instruments for all 10 biomarkers, R² values were 0.999 for 15/30 comparisons and R² ≥ 0.995 for 27/30 comparisons; CA9 showed the greatest variability (R² = 0.948–0.970). Standard curve slopes were statistically indistinguishable for all 10 biomarkers across analyzers. Conclusions: The Oncuria BC assay generates comprehensive urinary protein signatures useful for assisting BC diagnosis, predicting treatment response, and tracking disease progression and recurrence. The equivalent performance of the multiplex BC assay using three popular analyzers rationalizes test adoption by CLIA (Clinical Laboratory Improvement Amendments) clinical and research laboratories. Full article

Background and Aims: Preterm newborns are particularly susceptible to hypovitaminosis D, potentially impairing bone mineralization. In Thailand, data on its prevalence and standardized supplementation protocols remain limited. This study aimed to compare the efficacy of two vitamin D3 dosages (400 IU/day vs. 800 IU/day) in improving serum vitamin D concentrations and metabolic bone parameters in preterm newborns. Methods: A randomized controlled trial was conducted in preterm newborns born at ≤32 weeks’ gestation or with birth weight ≤1500 g. Preterm newborns were randomized to receive either 400 IU or 800 IU/day of vitamin D3. Serum 25-hydroxyvitamin D (25(OH)D) was measured using electrochemiluminescence immunoassay (ECLIA). Metabolic bone parameters—including calcium, phosphorus, alkaline phosphatase, and albumin—were assessed at baseline and again at six weeks of age. Results: Of the 38 enrolled infants, baseline 25(OH)D levels were comparable between groups (14.8 ± 4.8 ng/mL in the 800 IU/day group vs. 14.7 ± 6.9 ng/mL in the 400 IU/day group). At six weeks, the 800 IU group demonstrated significantly higher 25(OH)D levels (47.3 ± 21.0 ng/mL vs. 32.0 ± 14.2 ng/mL; p = 0.013), with a large effect size (Cohen’s d = 0.85) and the difference-in-differences of +15.7 ng/mL. The prevalence of hypovitaminosis D declined from 89% to 5% in the 800 IU/day group and from 74% to 32% in the 400 IU/day group (p = 0.036). No significant differences in metabolic bone parameters or signs of toxicity were observed. Conclusions: Vitamin D3 supplementation at 800 IU/day significantly improved vitamin D status and reduced hypovitaminosis D in preterm newborns, without observed toxicity. Full article

Background/Objective. Toxin-producing strains of Clostridioides difficile (C. diff) are the most commonly identified cause of healthcare-associated infection in the elderly. Risk factors include advanced age, hospitalization, prior or concomitant systemic antibacterial therapy, chemotherapy, and gastrointestinal surgery. Patients with unspecified and new-onset diarrhea with ≥3 unformed stools in 24 h are the target population for C. diff infection (CDI) testing. To present data on the risks of laboratory misdiagnosis in managing CDI. Materials. In two general hospitals, we examined 116 clinical stool specimens from hospitalized patients with acute diarrhea suspected of nosocomial or antibiotic-associated diarrhea (AAD) due to C. diff. Enzyme immunoassay (EIA) tests for the detection of C. diff toxins A (cdtA) and B (cdtB) in stool, automated CLIA assay for the detection of C. diff GDH antigen and qualitative determination of cdtA and B in human feces and anaerobic stool culture were applied for CDI laboratory diagnosis. MALDI-TOF (Bruker) was used to identify the presumptive anaerobic bacterial colonies. The following methods were used as confirmatory diagnostics: the LAMP method for the detection of Salmonella spp. and simultaneous detection of C. jejuni and C. coli, an E. coli Typing RT-PCR detection kit (ETEC, EHEC, STEC, EPEC, and EIEC), API 20E and aerobic stool culture methods. Results. A total of 40 toxigenic strains of C. diff were isolated from all 116 tested diarrheal stool samples, of which 38/40 produced toxin B and 2/40 strains were positive for both cdtA and cdtB. Of the stool samples positive for cdtA (6/50) and/or cdtB (44/50) by EIA, 33 were negative for C. diff culture but positive for the following diarrheal agents: Salmonella enterica subsp. arizonae (1/33, LAMP, culture, API 20E); C. jejuni (2/33, LAMP, culture, MALDI TOF); ETEC O142 (1/33), STEC O145 and O138 (2/33, E. coli RT-PCR detection kit, culture); C. perfringens (2/33, anaerobic culture, MALDI TOF); hypermycotic enterotoxigenic K. pneumonia (2/33) and enterotoxigenic P. mirabilis (2/33, culture; PCR encoding LT-toxin). Two of the sixty-six cdtB-positive samples (2/66) showed a similar misdiagnosis when analyzed using the CLIA method. However, the PCR analysis showed that they were cdtB-negative. In contrast, the LAMP method identified a positive result for C. jejuni in one sample, and another was STEC positive (stx1+/stx2+) by RT-PCR. We found an additional discrepancy in the CDI test results: EPEC O86 (RT-PCR eae+) was isolated from a fecal sample positive for GHA enzyme (CLIA) and negative for cdtA and cdtB (CLIA and PCR). However, the culture of C. diff was negative. These findings support the hypothesis that certain human bacterial pathogens that produce enterotoxins other than C. diff, as well as intestinal commensal microorganisms, including Klebsiella sp. and Proteus sp., contribute to false-positive EIA card tests for C. diff toxins A and B, which are the most widely used laboratory tests for CDI. Conclusions. CDI presents a significant challenge to clinical practice in terms of laboratory diagnostic management. It is recommended that toxin-only EIA tests should not be used as the sole diagnostic tool for CDI but should be limited to detecting toxins A and B. Accurate diagnosis of CDI requires a combination of laboratory diagnostic methods on which proper infection management depends. Full article

The rapid identification of stroke is critical to improving stroke patient outcomes. Existing protocols for assessing the risk of stroke are subjective and may be further complicated by nonspecific symptoms, increasing the risk of misdiagnosis. Neuron-specific enolase (NSE) has emerged as a promising stroke biomarker. However, current detection methods such as the electrochemiluminescence immunoassay (ECLIA) are time-consuming and costly. In this research, we developed an electrochemical biosensor for the rapid quantification of NSE in whole blood. Mouse stroke models were established, and blood samples collected were analyzed using both hospital-standard ECLIA as well as the biosensor. The biosensor limit of detection was 1.15 ng/mL. NSE measurements were highly correlated between the two methods and were obtained in 5 min using 20 μL of unprocessed whole blood samples. Notably, the biosensor could accurately quantify elevated blood NSE blood that was associated with more severe stroke. Our results demonstrate the utility of the proposed biosensor in pre-hospital settings. Combined with existing stroke assessment methods, the biosensor may enable emergency personnel to identify stroke risk with greater accuracy to optimize the chances of receiving necessary treatment within the effective window. Full article

Simple and accurate analysis of cancer-related biomarkers is very important for disease screening and auxiliary diagnosis. This study proposed a facile immunoassay that used gold nanostar-labeled rabbit anti-AFP as a capture antibody and gold nanoparticle-conjugated goat anti-rabbit IgG as an enhance antibody for the construction of a detection strategy for AFP analysis. Investigations indicated that the 50 nm diameter GNS-labeled capture antibody can specifically catch AFPs by direct detection profile or by further signal amplification through AuNP-tagged enhance antibody combination. Results showed that the developed method holds 8.6 ng/mL sensitivity, 20.0–110.0 ng/mL detection range, acceptable precision and fine accuracy, as well as favorable specificity. Results of application to real serum determination by the proposed method are highly related to those of the ECLIA method (correlation coefficient is 0.931). The proposed method has simple-operation merit and is very suitable for clinical screening of large-scale serum samples of cancers. Full article

Background/Objectives: Hepatitis E virus (HEV) is one of the leading causes of acute hepatitis, with immunosuppressed individuals, such as oncology patients, being particularly vulnerable to chronic infections that may progress to liver disease or fatal outcomes. Assay variability complicates HEV prevalence assessment in at-risk groups. This study aimed to compare the reliability and concordance of three HEV antibody assays—Wantai, Euroimmun, and Elecsys^®—in immunosuppressed oncology patients. Methods: In this prospective pilot study, serum samples were obtained from oncology patients between September 2020 and October 2021. Samples were collected both at baseline (treatment-naive) and during ongoing treatment. A healthy control group was retrospectively included for comparative analysis. Anti-HEV IgM and IgG antibodies were tested in all samples using enzyme-linked immunosorbent assays (Wantai, Euroimmun) and an electrochemiluminescence immunoassay (Elecsys^®). Demographic and clinical data, along with information on HEV risk factors, were extracted from medical records and patient questionnaires. Results: HEV IgM prevalence ranged from 0% (Wantai) to 6% (Elecsys^®), while IgG prevalence was 12% (Euroimmun), 38% (Wantai), and 53% (Elecsys^®). Concordance was poor, with Cohen’s Kappa values indicating slight to moderate agreement (κ = 0.000–0.553). Patients with hematological malignancies exhibited the highest IgG seroprevalence. Risk factor analysis revealed the highest association between HEV exposure and the consumption of undercooked pork or crop-based agriculture. Conclusions: Significant variability among HEV serological assays highlights the challenges of reliable HEV diagnostics in immunosuppressed oncology patients. Assay selection and improved testing strategies are critical for this high-risk group. Full article

Space flight exposes astronauts to stressors that alter the immune response, rendering them vulnerable to infections and diseases. In this study, we aimed to determine the levels of inflammasome activation in the brains of mice that were housed in the International Space Station (ISS) for 37 days. C57BL/6 mice were launched to the ISS as part of NASA’s Rodent Research 1 Mission on SpaceX-4 CRS-4 Dragon cargo spacecraft from 21 September 2014 to 25 October 2014. Dissected mouse brains from that mission were analyzed by immunoblotting of inflammasome signaling proteins and Electrochemiluminescence Immunoassay (ECLIA) for inflammatory cytokine levels. Our data indicate decreased inflammasome activation in the brains of mice that were housed in the ISS for 37 days when compared to the brains of mice that were maintained on the ground, and in mice corresponding to the baseline group that were sacrificed at the time of launching of SpaceX-4. Moreover, we did not detect any significant changes in the expression levels of the pro-inflammatory cytokines TNF-α, IL-2, IFN-γ, IL-5, IL-6, IL-12p70 and IL-10 between the ground control and the flight groups. Together, these studies suggest that spaceflight results in a decrease in the levels of innate immune signaling molecules that govern inflammasome signaling in the brain of mice. Full article

Rubella Virus, Cytomegalovirus (CMV), Herpes Simplex Virus-2 (HSV-2), Hepatitis B (HBV) and Hepatitis C virus (HCV) can cause serious fetal disease. The seropositivity rates of these agents vary among countries and geographic regions. This study aimed to analyze the prevalence rates and diagnostic methods used in studies investigating the seroprevalence of viral pathogens in the TORCH group among pregnant women in Turkey between 2005 and 2024. A systematic search was conducted using electronic databases between January 2005 and January 2024. A total of 60 studies meeting the inclusion criteria were included. Data quality control was assessed using the Joanna Briggs Institute guideline prevalence studies checklist. Heterogeneity was measured using the I-squared (I²) statistic in the Comprehensive Meta Analysis (CMA) program. The average seropositivity rates for Rubella, CMV, HSV-2, HBV and HCV in Turkey were determined as 91.18%, 94.81%, 35.52%, 1.66% and 0.25%, respectively. When the diagnostic methods were examined, it was determined that ELISA and ECLIA methods were used most frequently. The seropositivity of the agents did not show statistically significant differences according to the year periods, geographical regions and age of the patients (p > 0.05). The highest prevalence rates of Rubella and HSV-2 in pregnant women were reported in the Mediterranean region, the highest prevalence rates of CMV and HCV in the Southeastern Anatolia region and the highest seroprevalence of Anti HBs in the Marmara region. The results of this study support the necessity of increasing public awareness in the control of fetal infection caused by TORCH viral agents, prenatal screening, vaccination for Rubella and HBV and compliance with hygiene conditions for agents such as CMV, HSV-2 and HCV. The results of this study highlight the need to increase public awareness on prenatal screening for the control of fetal infection caused by all TORCH viral agents, vaccination for Rubella and HBV and compliance with hygiene conditions for agents such as CMV, HSV-2 and HCV. Full article

Vaginitis is a widespread issue for women worldwide, yet current diagnostic tools are lacking. Bacterial vaginosis (BV) is the most prevalent type of vaginitis, found in 10–50% of reproductive-aged women. Current diagnostic methods for BV rely on clinical criteria, microscopy, or the detection of a few microbes by qPCR. However, many vaginal infections lack a single etiological agent and are characterized by changes in the vaginal microbiome community structure (e.g., BV is defined as a loss of protective lactobacilli resulting in an overgrowth of anaerobic bacteria). Shotgun metagenomic sequencing provides a comprehensive view of all the organisms present in the vaginal microbiome (VMB), allowing for a better understanding of all potential etiologies. Here, we describe a robust VMB metagenomics sequencing test with a sensitivity of 93.1%, a specificity of 90%, a negative predictive value of 93.4%, and a positive predictive value of 89.6% certified by Clinical Laboratory Improvement Amendments (CLIA), the College of American Pathologist (CAP), and the Clinical Laboratory Evaluation Program (CLEP). We sequenced over 7000 human vaginal samples with this pipeline and described general findings and comparisons to US census data. Full article

(1) Background: In vitro diagnostic (IVD) tests are the main means of obtaining diagnostic information for clinical purposes. The electrochemiluminescence immunoassay (ECLIA) has become an important in vitro diagnostic technique. It has unique advantages and broad market prospects due to its sensitivity, detection limit, detection range and reagent stability. At present, there is a need to develop and optimize electrochemiluminescence immunoassay subsystems to achieve high-throughput outputs and accurate detection of instruments to promote their clinical application. (2) Methods: On the basis of the demand for clinical testing instruments with high detection accuracy and speed, this study constructed an electrochemiluminescence immunoassay system by designing magnetic separation modules, introducing PMT and optimizing the system timing regulation capability. (3) Results: The magnetic separation modules can increase the detection accuracy, the PMT system increases the detection sensitivity and optimized system timing can achieve maximum test output. Furthermore, this system performs well in terms of its linearity, detection limit, signal-to-noise ratio, precision and accuracy. (4) Conclusions: The electrochemiluminescence immunoassay system is capable of high throughput with good sensitivity and accuracy, meeting the basic requirements of clinical applications for detection capacity and output throughput. Full article

Growth hormone therapy (GHT) can improve growth velocity and final height, but can also accelerate the process of bone growth, which is related to structural bone modeling in both formation and resorption. This study evaluated the capacity of bone turnover markers to predict early growth response to one year of GHT in short stature children born small for gestational age (SGA). This study included 25 prepubertal children born SGA. We estimated P1NP (N-terminal procollagen type 1), CTX (C-terminal telopeptide of collagen type 1), P3NP (N-terminal procollagen type 3), NT-pro-CNP (amino-terminal C-type natriuretic peptide) and Ca-P metabolism using standard ECLIA (electrochemiluminescence), RIA (radioimmunoassay), and ELISA (enzyme-linked immunosorbent assay) methods. A statistically significant increase in bone resorption markers (CTX) was found at both 6 and 12 months. P1NP bone markers were increased at 6 months and after 12 months of therapy. The P3NP marker for collagen synthesis also increased after 12 months of therapy. We obtained significant increases in phosphorus levels at 6 and 12 months, and similar ALP (alkaline phosphatase) increases. We found a significant correlation between height (cm) and CTX after 6–12 months, as well as a P1NP/height (SD) correlation after 12 months. Calcium levels significantly correlated with height (SD) after 12 months. We found strong reactions of bone resorption and bone formation markers during growth hormone therapy, which may determine their selection as predictors of GHT outcome in children born SGA. However, the issue requires further research. Full article

Cystic fibrosis is caused by biallelic pathogenic variants in the CFTR gene, which contains a polymorphic (TG)_mT_n sequence (the “poly-T/TG tract”) in intron 9. While T₉ and T₇ alleles are benign, T₅ alleles with longer TG repeats, e.g., (TG)₁₂T₅ and (TG)₁₃T₅, are clinically significant. Thus, professional medical societies currently recommend reporting the TG repeat size when T₅ is detected. Sanger sequencing is a cost-effective method of genotyping the (TG)_mT_n tract; however, its polymorphic length substantially complicates data analysis. We developed CFTR-TIPS, a freely available web-based software tool that infers the (TG)_mT_n genotype from Sanger sequencing data. This tool detects the (TG)_mT_n tract in the chromatograms, quantifies goodness of fit with expected patterns, and visualizes the results in a graphical user interface. It is broadly compatible with any Sanger chromatogram that contains the (TG)_mT_n tract ± 15 bp. We evaluated CFTR-TIPS using 835 clinical samples previously analyzed in a CLIA-certified, CAP-accredited laboratory. When operated fully automatically, CFTR-TIPS achieved 99.8% concordance with our clinically validated manual workflow, while generally taking less than 10 s per sample. There were two discordant samples: one due to a co-occurring heterozygous duplication that confounded the tool and the other due to incomplete (TG)_mT_n tract detection in the reverse chromatogram. No clinically significant misclassifications were observed. CFTR-TIPS is a free, accurate, and rapid tool for CFTR (TG)_mT_n tract genotyping using cost-effective Sanger sequencing. This tool is suitable both for automated use and as an aid to manual review to enhance accuracy and reduce analysis time. Full article

“Background/Aim”: the current inability to diagnose Pancreatic Cancer Adenocarcinoma (PDAC) at an early stage strongly influences therapeutic strategies. Protein Induced by Vitamin K Absence (PIVKA II) showed an accurate diagnostic performance for PDAC. Since circulating PIVKA II has been recently associated with pancreatic origin cells with Vimentin, an epithelial-to-mesenchymal transition (EMT) early activation marker, the aim of this study was to investigate in vivo the combination between the two proteins. “Materials and Methods”: we assayed the presence of PIVKA II and Vimentin proteins by using different diagnostic methods. A total of 20 PDAC patients and 10 healthy donors were tested by Western Blot analysis; 74 PDAC patient and 46 healthy donors were assayed by ECLIA and Elisa. “Results”: Western Blot analysis showed the concomitant expression of PIVKA II and Vimentin in PDAC patient sera. Immunometric assay performed on a larger cohort of patients demonstrated that 72% of PIVKA II-positive PDAC patients were Vimentin-positive. Additionally, in a group of PDAC patients with PIVKA II levels ≥2070 ng/mL, the percentage of Vimentin-positive subjects reached 84%. “Conclusion“: the association between PIVKA II protein and the EMT suggests that this molecule could be considered a marker of the acquisition of an aggressive phenotype. Full article

The current diagnosis of attention deficit hyperactivity disorder (ADHD) is based on history, clinical observation, and behavioral tests. There is a high demand to find biomarkers for the diagnosis of ADHD. The aim of this study is to analyze the serum profiles of several biomarkers, including homocysteine (Hcy), vitamin B12, vitamin D, ferritin, and iron, in a cohort of 133 male subjects (6.5–12.5 years), including 67 individuals with an ADHD diagnosis based on DSM-V criteria and 66 age-matched healthy boys (healthy controls, HC). Assessments for ADHD included the Iowa Conners’ Teacher Rating Scale (CPRS) and the ADHDT test, as well as cognitive assessments using the Wechsler Intelligence Scale for Children-Revised (WISC-R) and the TROG-2 language comprehension test. Hcy and iron were quantified using spectrophotometry, while vitamin B12 and total 25-hydroxy vitamin D levels were determined using an electrochemiluminescence immunoassay (ECLIA) and ferritin was measured using a particle-enhanced immunoturbidimetric assay. The results showed significantly increased Hcy levels and decreased vitamin B12 levels in ADHD patients compared to HCs. Multiple logistic regression analysis indicated that Hcy is a potential prognostic indicator for ADHD. These results suggest that elevated homocysteine and decreased vitamin B12 may serve as markers for the diagnosis and prognosis of ADHD. Full article

The detection of anti-hepatitis E virus (HEV) antibodies contributes to the diagnosis of hepatitis E. The diagnostic suitability of two automated chemiluminescence immunoassays (CLIAs, LIAISON^® MUREX Anti-HEV IgG/Anti-HEV IgM test, DiaSorin) was assessed by comparison with the results of a combination of enzyme immunoassays and immunoblots (recomWell HEV IgG/IgM ELISA, recomLine HEV IgG/IgM, MIKROGEN). Samples with a deviating result were analyzed with the WANTAI ELISAs. Compared to the recomWell ELISAs, the Anti-HEV IgG CLIA had a percentage overall agreement (POA) of 100% (149/149; 95% CI: 97.5–100%) and the Anti-HEV IgM CLIA had a POA of 83.3% (85/102; 95% CI: 74.9–89.3%); considering the recomLine HEV IgM results, the POA was 71.7% (38/53; 95% CI: 58.4–82%). The WANTAI test confirmed 52.9% (9/17) of negative CLIA IgMs; HEV RNA was not detectable. Since acute infection with the Epstein–Barr virus (EBV) or human cytomegalovirus (CMV) may influence the results of other serological assays, HEV antibodies were examined in 17 EBV and 2 CMV patients: One had an isolated and probably unspecific HEV IgM in the CLIA, as HEV RNA was not detectable. Both CLIAs are well suited for HEV diagnostics, but isolated IgM should be confirmed. An acute EBV/CMV infection can influence HEV serodiagnostics. Full article