Electrochemiluminescence Assay Research Articles

Bone turnover, areal BMD, and bone microarchitecture by second-generation high-resolution peripheral quantitative computed tomography in transfusion-dependent thalassemia.

Thalassemic osteopathy includes low bone mass and impaired bone microarchitecture. We aimed to evaluate the prevalence and determinants of bone quantity (osteoporosis) and quality (microarchitecture) in a cohort of adult patients with transfusion-dependent thalassemia (TDT). Patients with TDT (n = 63) and age- and BMI-matched controls (n = 63) were recruited in the study. Areal bone mineral density (BMD) was measured using DXA Hologic scanner. P1NP and β-CTX were estimated by electrochemiluminescence assay. Bone geometry and volumetric BMD (vBMD) were estimated by second-generation high-resolution peripheral quantitative computed tomography. Bone turnover marker β-CTX was significantly lower in the TDT group, but there was no difference in P1NP levels. Low bone mass (Z ≤ -2) was present in greater proportion of patients both at lumbar spine (LS) (54 vs 0%; p = .001) and femoral neck (FN) (33 vs 8%; p = .001). Hypogonadism was associated with low BMD at FN (OR 10.0; 95% CI, 1.2-86; p = .01) and low hemoglobin with low BMD at LS (OR 1.58; 95% CI, 0.96-2.60; p = .07). The mean trabecular bone score was also significantly lower in patients compared with controls (1.261 ± 0.072 vs 1.389 ± 0.058). Total, cortical and trabecular vBMD were significantly lower in cases than controls. The trabecular number and cortical thickness were significantly lower and trabecular separation higher in cases than controls. Adults with TDT have significantly lower areal, cortical and trabecular vBMD. The bone microarchitecture is also significantly impaired in terms of lower number and wider spacing of trabeculae as well as lower cortical thickness and area at both radius and tibia.

Phase 2 Trial of Vosoritide Use in Patients with Hypochondroplasia: A Pharmacokinetic/ Pharmacodynamic Analysis.

Vosoritide is a C-type natriuretic peptide (CNP) analog that binds its receptor on chondrocytes, promoting growth by inhibiting the ERK1/2-MAPK pathway. We previously reported the results of a Phase II study in children with hypochondroplasia. Vosoritide led to an average increase in annualized growth velocity (AGV) of 1.81 cm/year and gain of 0.36 in height SD over 12 months. We present here the pharmacokinetic/ pharmacodynamic (PK/PD) data from this study and examine the correlations between these parameters and growth outcomes. We conducted a Phase II trial of daily subcutaneous vosoritide (15 mcg/kg/day) in 24 prepubertal subjects with hypochondroplasia (12 females, mean age 5.9+/-2.3 years, mean height -3.29+0.68 SD). Plasma vosoritide levels were assayed using an electrochemiluminescence assay. Pharmacodynamic markers including serum collagen X biomarker (CXM) and urine cGMP production were measured at Day 1, Month 6 and Month 12 visits. Pearson correlations and regression analyses were performed between PK and PD parameters and growth outcomes. Vosoritide PK parameters were similar to those previously reported in patients with achondroplasia. CXM levels increased from a baseline mean of 22.5±6.5 to 41.6±15.9 ng/ml after 12 months of treatment (p < 0.0001). Urine cGMP increased within 1 hour and peaked at 2 hours after injection. The mean AUC for cGMP production was not significantly different at each study visit. The maximum change in cGMP AUC correlated with PK AUC ((r=0.46, p=0.0001). However, drug exposure, as measured by average PK AUC, did not correlate with any growth outcome. CXM levels correlated with the prior 6-month interval height velocity (partial correlation coefficient=0.40, p=0.0048). However, change in CXM did not correlate with change in height velocity or change in height SD during treatment. Vosoritide treatment showed improvement in AGV and height SD in children with hypochondroplasia. PK analysis indicates that drug exposure was correlated to global CNP activity as measured by urine cGMP but did not correlate with growth outcomes. More studies are needed to identify specific patient characteristics that can predict response to therapy and clinical outcomes.

Highly Sensitive Electrochemiluminescence Biosensing Method for SARS-CoV-2 N Protein Incorporating the Micelle Probes of Quantum Dots and Dibenzoyl Peroxide Using the Screen-Printed Carbon Electrode Modified with a Carboxyl-Functionalized Graphene.

Obtaining stable electrochemiluminescence (ECL) emissions from a hydrophobic luminophore in aqueous solutions and designing a method without the use of an exogenous coreactant are promising for ECL biosensing. Here, a highly sensitive signal-on ECL immunoassay for the SARS-CoV-2 N protein was developed using micelles as an ECL tag. The micelles were prepared by coencapsulating the luminophore hydrophobic CdSe/ZnS quantum dots and coreactant dibenzoyl peroxide within the hydrophobic core of micelles. The ECL probe was obtained by covalently bonding a SARS-CoV-2 N protein-binding aptamer onto the micelle surface. The construction of the immunosensor was initiated by the immobilization of the anti-SARS-CoV-2 N protein antibody onto the screen-printed carbon electrode (SPCE) with a -COOH-functionalized surface. The surface functionalization of SPCEs was achieved through paste-exfoliated graphene, which was modified with a -COOH group through supramolecular-covalent scaffolds on SPCE. Upon achieving sandwich complexes on the immunosensor, an efficient ECL signal response at -1.4 V versus Ag/AgCl was obtained in phosphate buffer solution. The ECL assay was used for the sensitive determination of SARS-CoV-2 N protein with the linear range from 0.01 to 50 ng mL-1, and the detection limit was 3.0 pg mL-1. The immunosensor showed good reproducibility and stability, and the ECL immunoassay was used to determine the SARS-CoV-2 N protein in serum samples. The proposed approach to obtain micelles is versatile for the preparation of stable ECL luminophores by using hydrophobic materials, and the strategy provides an alternative for ECL bioassays based on the coreactant route.

Association of serum interferon alpha-2a levels with disease severity and prognosis in systemic sclerosis.

To determine serum type I interferon IFN-α2a concentrations in systemic sclerosis (SSc) patients, explore its association with cytokine/chemokine expressions, and evaluate correlation with the phenotype including the predictive value for interstitial lung disease (ILD) progression. Serum samples were obtained from 200 SSc patients and 29 healthy controls. IFN-α2a levels were measured by ultrasensitive electrochemiluminescence assay. Pro-inflammatory and chemokine panels were determined by Luminex® Discovery Assay multiplex kit. Baseline SSc disease characteristics were recorded together with longitudinal data for determining ILD progression after 2 years. IFN-α2a concentrations were higher in SSc patients compared with controls, although not reaching significance (means ± SD of 49.20 ± 156.8 pg/ml vs 9.606 ± 4.399 pg/ml, respectively; (p = 0.158)). Using the cut-off of 15.9 pg/ml, we identified 62 patients as having a type 1 (T1) IFN signature in their circulation. Patients with an IFN signature had significantly higher levels of chemokines (CCL8, CCL19, CXCL10, CXCL11), and the cytokine IL-1α compared with those without an IFN signature. IFN-α2a concentrations strongly correlated with a T1 IFN related chemokine score supporting activation of this pathway. Phenotyping association queries revealed association between IFN values and both skin and ILD involvements at baseline. Longitudinal data did not identify IFN as a predictive marker for ILD progression. Using serum determinations, the activation of the T1 IFN pathway showed strong correlations with inflammatory mediators and associations with clinical manifestations, especially skin fibrosis and ILD in SSc patients. However, activated IFN pathway was not predictive of ILD progression.

Monitoring Estradiol and Progesterone Content of Saanen Goats During Pregnancy

Saanen is a specialized dairy goat breed originating from Switzerland, imported and encouraged to be raised in Vietnam. This article presents the results of a survey of estradiol and progesterone levels during pregnancy in Saanen goats at the Binh Duong Large Livestock Research and Development Center. After monitoring mating, goats were sampled for blood every 7 days until birth, at 6-8 am. Hormone levels were determined by electrochemiluminescence assay, with the Cobas E601 immunoassay system. The results showed that after 14 days of conception, progesterone levels gradually increased (3.21±0.36 ng/ml) and shifted to higher levels, peaking on day 70 of pregnancy (12.28±0.52 ng/ml) (in the 1-2 years old group); corresponding values were 3.18±0.75-11.88±0.92 ng/ml (in the 3-4 years old group). The high value range was maintained throughout pregnancy and decreased rapidly in the 14-21 days before birth in both age groups. Estradiol, maintained at low concentrations from 0.73±0.35 to 2.65±1.25 pg/ml (in the 1-2 years old group); from 0.98±0.48 to 3.27±1.95 pg/ml (in the 3-4 years old group) in the first 21 days of pregnancy and then gradually increased to above 10 pg/ml (in both age groups) on day 28. Estradiol continued to increase and reached a maximum level (205.48±32.81 pg/ml) in group 1 on day 126 and had an average value below 10 pg/ml within 1-2 days after giving birth. The results of this study help diagnose pregnancy, predict gestational age for effective reproductive management, and serve as a reliable reference when using assisted reproductive tools and strategies to improve reproductive performance in goats.

The Endogenous Expression of BMI1 in Adult Human Eyes.

BMI1, also known as B lymphoma Mo-MLV insertion region 1, is a protein in the Polycomb group that is implicated in various cellular processes, including stem cell self-renewal and the regulation of cellular senescence. BMI1 plays a role in the regulation of retinal progenitor cells and the renewal of adult neuronal cells. However, the presence, location, and quantification of BMI1 in the adult human eye have never previously been reported. In this study, we collected 45 frozen globes from eye banks, and ocular tissues were dissected. Protein was quantified by utilizing a custom electrochemiluminescence (ECL) assay developed to quantify the BMI1 protein. BMI1 was found in all ocular tissues at the following levels: the retina (1483.6 ± 191.7 pg/mL) and the RPE (296.4 ± 78.1 pg/mL). BMI1 expression was noted ubiquitously in the GCL (ganglion cell layer), the INL (inner nuclear layer), the ONL (outer nuclear layer), and the RPE (retinal pigment epithelium) via immunofluorescence, with higher levels in the inner than in the outer retinal layers and the RPE. These data confirm that BMI1 is expressed in the human retina. Further studies will illuminate the role that BMI1 plays in ocular cells. BMI1 levels are lower in aged retinas, possibly reflecting changes in retinal somatic and stem cell maintenance and disease susceptibility.

The fetal origins of metabolic health: exploring the association between newborn biological age and metabolism hormones in childhood

BackgroundTelomere length (TL), mitochondrial DNA copy number (mtDNAcn), and DNA methylation age (DNAmAge) are common aging biomarkers. However, research on the associations between these three markers at birth and subsequent metabolic status was limited. This study aimed to evaluate the association between TL, mtDNAcn, and DNAmAge in newborns and the variation in metabolic hormones of children at 3 years old.MethodsThis research involved 895 mother–child pairs from a birth cohort in China, with TL and mtDNAcn measured using quantitative real-time PCR, DNA methylation (DNAm) assessed using Infinium MethylationEPIC Beadchip, and DNAm age (DNAmAge) determined using Horvath’s epigenetic clock. Insulin and leptin levels were measured via electrochemiluminescence assay. Multivariable adjusted linear regression and restricted cubic spline (RCS) analysis were utilized to examine the association between aging markers and metabolic hormones.ResultsThe linear regression analysis indicated the percentage change of metabolism hormones for per doubling of aging biomarkers alterations and found significant associations between DNAmAge and insulin levels (adjusted percent change (95% CI), − 13.22 (− 23.21 to − 1.94)), TL and leptin levels (adjusted percent change (95% CI), 15.32 (1.32 to 31.24)), and mtDNAcn and leptin levels (adjusted percent change (95% CI), − 14.13 (− 21.59 to − 5.95)). The RCS analysis revealed significant non-linear associations between TL (Ln transformed) and insulin (Ln transformed) (P = 0.024 for nonlinearity), as well as DNAmAge (Ln transformed) and leptin (Ln transformed) (P = 0.043 for nonlinearity). Specifically, for TL and insulin, a positive association was observed when TL (Ln transformed) was less than − 0.05, which transitioned to an inverse association when TL (Ln transformed) was greater than − 0.05. Regarding DNAmAge and leptin, there was a sharp decline when DNAmAge (Ln transformed) was less than − 1.35, followed by a plateau between − 1.35 and − 0.67 and then a further decline when DNAmAge (Ln transformed) was greater than − 0.67.ConclusionsIn this prospective birth cohort study, variation in metabolic hormones of children at 3 years old was associated with TL, mtDNAcn, and DNAmAge at birth. These findings suggested that TL, mtDNAcn, and DNAmAge might play a role in the biological programming of metabolic health from birth.

B-313 Assessment of β2-microglobulin concentration in pleural effusions

Abstract Background β2-microglobulin (β2-m) is a biomarker used in serum for different types of pathologies. It has been described that lymphocyte activation processes us tuberculosis infections, AIDS virus or cytomegalovirus infections, can produce an increase in the concentration of this marker. As a tumor marker is useful in lymphomas diagnosis. In the bibliography has reported that processes involving renal failure lead to a significant increase in its concentration. In patients with renal failure and treated with hemodialysis, have been described medians twenty times the upper reference limit. The objective is to assess whether the concentration of (β2-m) in pleural fluid is affected by the same factors as in serum. Methods We analyzed 303 pleural effusions (PE) samples from undiagnosed patients with pleural effusion. β2-microglobulin levels were prospectively measured by electrochemiluminescence (ECLIA) assays (Cobas c702, Roche Diagnostics®). Glomerular filtration rate (GFR) was calculated by CKD-EPI equation. The diagnostic performance of β2-microglobulin was evaluated by receiver operating characteristic (ROC) curve analysis. Statistical analyses were performed using SPSS software. Results Pleural fluid of 303 patients (104 female, 204 male) were collected. The median age was 72 ± 10,6 years. For all samples β2-m median concentration was 5,85 mg/L (IQR:3.4 -6.45).When we compared all results according to kidney failure values; GFR >60mL/min β2-m median concentration was 4,12mg/L and if GFR <60 mL/min, β2-m was 8,62g/L significant difference have found (p<0,001). Comparing results between malignant pleural effusions (MPE) (n=96) and benign pleural effusions (BPE) (n=207), MPE median concentration was 5.38 mg/L and 6.07mg/L significant difference (p<0,001). Benign pathologies associated with pleural effusions, tuberculosis samples had the highest concentration of β2-m. Tuberculosis samples (n=6) had a normal distribution, obtaining a value [mean (SD)] of 7.28 mg/mL (2,13) mg/mL. For tuberculosis samples and patients without renal failure (n=5) (GFR>60 mL/min), the area under the ROC curve was 0.98 (95% CI: 0.88-0.98) (p<0.005). For β2-m cut-off point was 5.150 mg/L, a sensitivity of 100%, specificity of 16%, positive predictive value (PPV) of 14% and negative predictive value (NPV) of 100% were obtained. Patients with lymphomas present the highest concentration in the group of malignant pleural effusions, but further investigations could not be performed due to the small sample size. Conclusions Based on the results obtained from our present study, it can be concluded that β2-microglobulin marker in pleural effusions has significant association with glomerular filtration rate, as well as occurs in serum. Higher concentrations were found in patients with GFR>60 mL/min and tuberculosis. Using 5,15 mg/L cut-off point, 100% of NPV was obtained.

A-242 Assessing the efficacy of the one-step diagnostic algorithm for Hepatitis C

Abstract Background Hepatitis C virus (HCV) infection can be asymptomatic and may go into spontaneous remission. However, in 60-80% of cases, it becomes chronic and progresses into cirrhosis or hepatocarcinoma. The virus is mainly transmitted parenterally, although cases of sexual and transplacental transmission have also been found. Since the development of direct-acting antiviral (DAA) therapy that can achieve a complete cure in 95% of patients, eradication strategies based on rapid diagnosis and treatment have been promoted. Therefore, the World Health Organization (WHO) has set a goal to eliminate HCV infection as public health threat by 2030. This is a retrospective study on the usefulness and effectiveness of the one-step HCV diagnostic algorithm application from May 2022 to December 2023 in our hospital. Methods The current HCV testing algorithm consists in a reflex HCV RNA testing, in which anti-HCV antibody-positive samples are automatically referred for HCV RNA testing on the same specimen. We used a serological electrochemiluminescence (ECLIA) assay for the detection of anti-HCV antibodies (Cobas e801, Roche Diagnostics®) in serum samples; and then a confirmation of HCV viremia by nucleic acid testing of HCV RNA in plasma samples (Cobas 4800, Roche Diagnostics®). Results A total of 22713 HCV serologies were performed with a total of 221 positive results (1%). Of the HCV cases, 21 patients had a newly diagnosis of active infection (10%) for whom we recommended derivation to the Digestive Service to initiate elimination treatment. 71% were men, with a mean age of 53 years and a standard deviation of 17. The mean age of women was 56 years with a standard deviation of 18. Coinfection with HIV or hepatitis B virus (HBV) was also assessed. Patients with HIV-HCV coinfection represented 9% of cases and were on average 12 years younger than HCV-monoinfected individuals. We did not identify any case of HBV-HCV co-infection. Conclusions The pattern we observed in terms of incidence by age and sex is similar to that reported in the literature, as well as the incidence of new HCV cases. One-step diagnosis enables the laboratory to identify the need for referring to a specialist, reducing the time to treatment for new and known but untreated cases. Therefore, global access to HCV diagnostics will be a keystone to success.

A-074 A Laboratory Developed Serum Anti-Mullerian Hormone Electrochemiluminescent Immunoassay: Performance and Retrospective Analysis of Over 2.2 Million Tests in Females and Males Across Lifespan

Abstract Background The quantification of serum Anti-Mullerian hormone (AMH), a glycoprotein dimer produced by granulosa cells of the ovarian follicle in females and by Sertoli cells of the testes in males, has clinical utility across lifespan in both females and males. AMH is most frequently ordered to assess fertility in females of reproductive age, where it correlates with ovarian reserve. The clinical applications of serum AMH beyond ovarian reserve assessment, especially in males, pediatric, and postmenopausal populations, include the investigation of peri-menopause transition, delayed or precocious onset of puberty, differential diagnosis of disorders of sexual development, the evaluation of male gonadal function, and as an ovarian tumor marker. The objective was to establish and validate a highly sensitive assay to quantify AMH and to use our database of more than 2 million tests to understand the clinical utility of measuring AMH across sex and lifespan. Methods A validated two-step electrochemiluminescence assay was used for the quantification of serum AMH. The analytical performance of the assay was assessed, including accuracy, precision, and dilutional linearity. ICD-10 codes ordered between 2012 and 2023 were retrospectively analyzed for each subgroup based on age and sex to determine test usage. The analysis includes results from 2,229,845 female and 10,362 male patients across lifespan. Results The AMH analytical measurement range (AMR) was determined to be 0.015 - 15.0 ng/mL with a maximum reportable concentration of 960 ng/mL when run on dilution. Both inter- and intra-assay standard accuracy and precision were ≤ ±15.4% bias and ≤ 11.0% CV for all levels. Inter- and intra-assay sample precision was ≤ 9.6% CV across the AMR. Samples diluted linearly and recovered within 80 - 120% of the expected values when diluted up to 64X. Analytical interference caused by icterus, lipemia, or hemolysis was not observed. The most common ICD-10 associated with AMH orders was for fertility testing in females of reproductive age. AMH was also ordered in females for evaluation of ovarian cancer, menstrual disorders, and PCOS. In males, common ICD-10 codes associated with AMH include delayed puberty, hypoplasia of the penis, fertility testing, hypogonadism, and childhood short stature. Conclusions This study demonstrates a highly sensitive assay for the quantification of serum AMH with clinical utility across sex and lifespan. Using retrospective data, we determined that in addition to being used primarily in fertility testing, AMH is also commonly used to evaluate and manage other endocrine disorders and malignancies, including disorders of sexual development, disorders of puberty, PCOS, male gonadal dysfunction, and granulosa cell tumors. This assay can aid in defining the clinically relevant age and sex-specific levels of AHM in non-fertility-related conditions.

Seroprevalence of Neonatal Herpes Simplex Virus Infection at A Tertiary Teaching Hospital in Malaysia

INTRODUCTION: Neonatal herpes simplex virus (HSV) infection is generally infrequent, despite being a life-threatening illness. Knowledge of its prevalence is limited in Malaysia since most cases are asymptomatic infections and only limited routine neonatal HSV screening is conducted. This study therefore provides a comprehensive investigation of the seroprevalence of HSV-1 and HSV-2 in neonates. MATERIALS AND METHODS: Serological screening for HSV-1 IgG and HSV-2 IgG antibody tests using the Electrochemiluminescence assay was performed on serum samples of 215 neonates delivered from January until December 2022 at Hospital Universiti Sains Malaysia. RESULTS: Of the neonates, 54.4% were found to be HSV-1 positive, while 4.2% were HSV-2 positive. All the HSV-2 neonates were co-infected with HSV-1. Newborns aged 0–10 days were the most infected group by HSV-1 (92.3%) and HSV-2 (55.6%). The most reported clinical presentation was small gestational age (SGA) (60%). Microcephaly and macrocephaly were observed in one neonate each. The clinical presentations of reactive HSV-1 and HSV-2 cases revealed the presence of fever with rash in both cases. CONCLUSION: The high seroprevalence of HSV-1 is alarming. It is hoped that these data will support the advocacy of screening women for HSV before or during pregnancy as a precautionary approach to reducing the risk of vertical transmission.

Molecular Mechanisms of Neurofilament Alterations and its Application in Assessing Neurodegenerative Disorders.

Neurofilaments are intermediate filaments present in neurons. These provide structural support and maintain the size and shape of the neurons. Dysregulation, mutation, and aggregation of neurofilaments raise the levels of these proteins in the blood and cerebrospinal fluid, which are characteristic features of axonal damage and certain rare neurological diseases, such as Giant axonal neuropathy and Charcot-Mare-Tooth disease. Understanding the structure, dynamics, and function of neurofilaments has been greatly enhanced by a diverse range of biochemical and preclinical investigations conducted over more than four decades. Recently, there has been a resurgence of interest in post-translational modifications of neurofilaments, such as phosphorylation, aggregation, mutation, oxidation, etc. Over the past twenty years, several rare disorders have been studied from structural alterations of neurofilaments. These disorders are monitored by fluid biomarkers such as neurofilament light chains. Currently, there are many tools, such as Enzyme-Linked Immunosorbent Assay, Electrochemiluminescence Assay, Single-Molecule Array, Western/immunoblotting, etc., in use to assess the neurofilament proteins in Blood and CSF. However, all these techniques utilize expensive, non-specific, or antibody-based methods, which make them unsuitable for routine screening of neurodegenerative disorders. This provides room to search for newer sensitive, cost-effective, point-of-care tools for rapid screening of the disease. For a long time, the molecular mechanisms of neurofilaments have been poorly understood due to insufficient research attempts, and a deeper understanding of them remains elusive. Therefore, this review aims to highlight the available literature on molecular mechanisms of neurofilaments and the function of neurofilaments in axonal transport, axonal conduction, axonal growth, and neurofilament aggregation, respectively. Further, this review discusses the role of neurofilaments as potential biomarkers for the identification of several neurodegenerative diseases in clinical laboratory practice.

Serum concentrations of chromogranin A, serotonin and natriuretic peptide are decreased in patients with neuroendocrine tumors with overweight and obesity compared to healthy donors

Background: Neuroendocrine tumors (NETs) are a group of neoplasms, in which circulating biomarkers chromogranin A (CgA), serotonin, and N-terminal brain natriuretic pro-peptide (NT-proBNP), used as a marker of carcinoid heart disease, are especially important in the diagnosis and monitoring. A number of pre-analytical factors, including patients' body mass index (BMI), have an impact on NETs biomarker concentrations. Aim: To perform a comparative analysis of serum concentrations of CgA, serotonin and NT-proBNP in patients with NETs of various locations with normal body weight and with overweight or obesity. Methods: This cross-sectional study included 94 patients with NETs of various locations and 78 provisionally healthy individuals without cancer and cardiovascular disorders, matched by gender and age to the patients. Serum biochemical markers were measured before a course of chemotherapy/biotherapy or surgery with the use of standardized enzyme-linked immunosorbent assays Chromogranin A NEOELISA (Eurodiagnostica), Serotonin ELISA (IBL), and the electrochemiluminescent assay (Cobas e601 analyzer, Roche). All NETs patients were divided into 2 subgroups: 46 patients with normal bodyweight (BMI 18 to 24.9) and 48 patients with overweight/obesity (BMI ≥ 25). Results: The median BMI in the NET patients did not vary depending on their tumor extension, functional activity and malignancy grade. The median concentrations of CgA (150 ng/ml), serotonin (188 ng/ml) and NT-proBNP (117 pg/ml) in the NET patients with obesity and overweight were significantly lower (p = 0.008, p = 0.005 and p = 0.012, respectively) than in the NET patients with normal body weight (769 ng/ml, 704 ng/ml and 197 pg/ml, respectively). In the control group, significantly (p = 0.0001) lower levels in the subgroup with BMI ≥ 25 were obtained only for serotonin. The ROC analysis showed a decrease of the diagnostic efficiency of NET biomarkers in obesity: in the normal BMI group AUC (CgA) was 0.87 and AUC (serotonin) 0.78, whereas in those with BMI ≥ 25, the AUC (CgA) and AUC (serotonin) were 0.81 and 0.62, respectively. Conclusion: Obesity may make it difficult to identify biochemical markers of NETs – CgA, serotonin and NT-proBNP.

Multifunctional spatial-potential resolved electrochemiluminescence biosensor for dual-channel simultaneous detection of hepatitis A virus and hepatitis C virus DNA

Highly sensitive simultaneous detection of dual-target is one of the main research directions of electrochemiluminescence (ECL), but the traditional three-electrode system makes it difficult to realize this aim. This work designed a multifunctional spatial-potential resolved ECL biosensor for dual-channel simultaneous detection of hepatitis A virus (HAV) and hepatitis C virus (HCV) DNA. Self-luminescent dual-ligand metal organic framework (D-MOF) and cerium MOF (Ce-MOF) were modified on two parts of the ITO electrode to achieve dual-channel simultaneous detection. One ligand of D-MOF acted as the luminophore and the other as the co-reactant, so D-MOF could emit excellent positive potential ECL signal without requiring external co-reactants. In channel A, the Ag nanoparticles (Ag NPs) quenching probes were introduced to D-MOF through hybridization of HAV with “Y” type DNA. Positive potential signal of D-MOF was decreased for HAV DNA detection based on resonance energy transfer (RET). In channel B, the CdSe quantum dots (QDs) were introduced to Ce-MOF through hybridization of HCV with “Y” type DNA. Due to the excellent ECL performance of CdSe QDs, negative potential signal was increased for HCV DNA detection. In this work, an ingenious spatial-potential resolved ECL biosensor was developed for simultaneous detection of dual-target by simple sealing film isolation, which opened a new strategy for dual-target detection in general ECL assays with three-electrode system.

Selective detection of ketamine in human urine samples by electrochemiluminescence sensor based on conductive molecular imprinted polypyrrole: A hydrogen peroxide‐free electrochemiluminescence assay

AbstractA new sensor was fabricated for the determination of ketamine (Ket) in urine samples based on enhanced electrochemiluminescence (EECL) behavior of luminol on a carbon paste electrode (CPE) modified with the conductive molecularly imprinted polymer (CMIP) film. The FESEM technique was applied to demonstrate the rod shape morphology of CMIP. Unlike the common luminol system, EECL signal of luminol no need to addition of hydrogen peroxide and it might find application in efforts to avoid instability of hydrogen peroxide or its interference with the analytes of interest. The EECL emission has been optimized by an investigation of the applied potentials, electrochemical polymerization conditions, template and modifier concentrations, and the pH solution. Under the optimum conditions, the EECL signal depended linearly on the Ket concentration within a range from 0.1 to 1.25 μM with a limit of detection (LOD) of 0.08 μM. The sensor exhibited very good recovery percentages ranging from 92% to 101% in urine samples, while precisions were ≤3%. These results suggested that the proposed sensor would be applicable for analysis Ket in complex matrixes without interfering of other drugs (anesthetics) and thus has the potential to be used in clinical application.

Longitudinal analysis at pre- and post-flare of T peripheral helper and T follicular helper subsets in patients with systemic lupus erythematosus

ObjectiveWe focused to analyze the time-course changes at pre- and post-flare of T peripheral helper (Tph) cells and circulating T follicular helper (Tfh) cells in the blood of patients with systemic lupus erythematosus (SLE) with lupus low disease activity state (LLDAS) before flare. MethodsThis study included inactive (n = 29) and active (n = 55) patients with SLE. Tph subsets, Tfh subsets, CD11chi B cells, and plasma cells in the blood were determined by flow cytometry. The blood levels of cytokines including interferons (IFNs) were measured by electrochemiluminescence assay or cytokine beads array. ResultsActive SLE patients exhibited the increased frequency of Tph1, Tph2, Tfh1, and Tfh2 subsets when compared to inactive patients, but no clear changes in the other subsets. During the treatment with medications, Tph1, Tph2, and Tfh2 subsets were significantly reduced along with disease activity and Tph1 and Tph2 subsets were positively correlated with SLE disease activity index (SLEDAI). The time course analysis of patients at pre- and post-flare revealed that in the patients at LLDAS before flare, Tph subsets and Tfh subsets were relatively low levels. At the flare, Tph cells, particularly Tph1 and Tph2 subsets, were increased and correlated with SLEDAI. Furthermore, the blood levels of IFN-α2a, IFN-γ, and IFN-λ1 were low in the patients with LLDAS before flare but these IFNs, particularly IFN-λ1, were increased along with flare. ConclusionIncreased frequency of Tph1 and Tph2 subsets and elevated levels of serum IFN-λ1 are presumably critical for triggering of flare in SLE.

Impact of nucleic acid testing in double screening of blood donations in Northern India: A single-center experience

Objectives: The aim of our study is to assess the ability of nucleic acid testing (NAT) to detect hepatitis B virus (HBV), hepatitis C virus (HCV), and human immunodeficiency virus (HIV) from donor blood samples which were declared sero-non-reactive. Materials and Methods: The whole blood donations were collected over the 6-year period from 2017 to 2023 which were initially screened for hepatitis B surface antigen, anti-HCV, anti-HIV-1/2, and p24 antigen and syphilis by electrochemiluminescence assays by Roche Diagnostics, Germany and malaria by Pan/Pf rapid test. Donations were declared nonreactive by these tests which were further subjected to NAT testing using minipool method. It was done in pool of 6 using cobas Taq Screen MPX version 2.0 on the cobas s201 platform. The NAT yields were quantitated for viral loads and followed up by serology. Statistical analysis: The data obtained was entered in Microsoft Excel and analyzed using descriptive statistics using the Statistical Package for the Social Science 21 version. Results: Out of 152,575 donations, 149,304 were sero-non-reactive and screened by NAT. Out of 82, 45 were reactive for HBV deoxyribonucleic acid and 37 for HCV ribonucleic acid. The NAT yield was 1:1831 overall, 1:3337 for HBV and 1:4059 for HCV. Viral load was quantitated in 65/82 NAT yields (35 HBV and 30 HCV). The viral loads of HBV samples were <20 IU/mL for 19/35 samples and the HCV sample viral loads ranged from 25.3 to 9.3 × 106 IU/mL. Eleven NAT-yield donors (4 HBV, 7 HCV) reported for follow-up showed sero-conversion between 90 and 210 days after NAT screening. Conclusions: This study confirmed the benefit of a highly sensitive minipool NAT (MP-NAT) in interdicting infectious donations undetected by conventional screening and illustrates the detection of donations with low viral loads using the MP-NAT polymerase chain reaction.

GDF-15 is associated with sarcopenia and frailty in acutely admitted older medical patients.

Growth differentiation factor-15 (GDF-15) has been associated with senescence, lower muscle strength, and physical performance in healthy older people. Still, it is not clear whether GDF-15 can be utilized as a biomarker of sarcopenia and frailty in the early stages of hospitalization. We investigated the association of plasma GDF-15 with sarcopenia and frailty in older, acutely admitted medical patients. The present study is based on secondary analyses of cross-sectional data from the Copenhagen PROTECT study, a prospective cohort study including 1071 patients ≥65years of age admitted to the acute medical ward at Copenhagen University Hospital, Bispebjerg, Denmark. Muscle strength was assessed using handgrip strength, and lean mass was assessed using direct segmental multifrequency bioelectrical impedance analyses and used to clarify the potential presence of sarcopenia defined according to guidelines from the European Working Group on Sarcopenia in Older People. Frailty was evaluated using the Clinical Frailty Scale. Plasma GDF-15 was measured using electrochemiluminescence assays from Meso Scale Discovery (MSD, Rockville, MD, USA). We included 1036 patients with completed blood samples (mean age 78.9±7.8years, 53% female). The median concentration of GDF-15 was 2669.3pg/mL. Systemic GDF-15 was significantly higher in patients with either sarcopenia (P<0.01) or frailty (P<0.001) compared with patients without the conditions. Optimum cut-off points of GDF-15 relating to sarcopenia and frailty were 1541 and 2166pg/mL, respectively. Systemic GDF-15 was higher in acutely admitted older medical patients with sarcopenia and frailty compared with patients without. The present study defined the optimum cut-off for GDF-15, related to the presence of sarcopenia and frailty, respectively. When elevated above the derived cutoffs, GDF-15 was strongly associated with frailty and sarcopenia in both crude and fully adjusted models.

A randomized phase 1/2a trial of ExPEC10V vaccine in adults with a history of UTI

The safety, reactogenicity, and immunogenicity of 3 doses of ExPEC10V (VAC52416), a vaccine candidate to prevent invasive Escherichia coli disease, were assessed in a phase 1/2a study (NCT03819049). In Cohort 1, ExPEC10V was well tolerated; the high dose was selected as optimal and further characterized in Cohort 2. Cohort 2 comprised a maximum 28-day screening, vaccination (Day 1), double-blind 181-day follow-up, and open-label long-term follow-up until Year 1. Healthy participants (≥60 years) with a history of urinary tract infection (UTI) within 5 years were randomized to receive ExPEC10V or placebo. The primary endpoint evaluated the safety and reactogenicity of ExPEC10V (solicited local and systemic AEs [until Day 15]; unsolicited AEs [until Day 30], SAEs [until Day 181], and immunogenicity [Day 30]) via multiplex electrochemiluminescent (ECL) and multiplex opsonophagocytic assay (MOPA). 416 participants (ExPEC10V, n = 278; placebo, n = 138) were included (mean age [SD], 68.8 [6.52] years; female, 79.6%; White, 96.1%). The incidence of solicited AEs was higher with ExPEC10V (local, 50.0% [n = 139]; systemic, 50.0% [n = 139]) than placebo (15.9% [n = 22]; 38.4% [n = 53]); rates of unsolicited AEs were comparable (ExPEC10V, 28.4% [n = 79]; placebo, 26.1% [n = 36]). No vaccine-related SAEs or deaths were reported. ExPEC10V elicited a robust antibody-mediated immunogenic response across all serotypes with ECL (Day 30 geometric mean fold increase, 2.33–8.18) and demonstrated functional opsonophagocytic killing activity across all measured serotypes (Day 30 geometric mean fold increase, 1.81–9.68). ExPEC10V exhibited an acceptable safety profile and a robust vaccine-induced functional immunogenic response in participants with a history of UTI. Clinical trial registration details: https://clinicaltrials.gov/study/NCT03819049.

Circulating kidney injury molecule-1 (KIM-1) biomarker analysis in IMmotion010: A randomized phase 3 study of adjuvant (adj) atezolizumab (atezo) vs placebo (pbo) in patients (pts) with renal cell carcinoma (RCC) at increased risk of recurrence after resection.

4506 Background: In IMmotion010, adj atezo did not prolong investigator-assessed disease-free survival (DFS; primary endpoint) vs pbo after resection in pts with RCC (HR: 0.93, 95% CI: 0.75, 1.15; P=0.50; Pal Lancet 2023). This exploratory analysis of circulating protein biomarkers was performed to identify high-risk pts with minimal residual disease (MRD) who may show differential benefit from treatment (tx) with atezo. Methods: Pts with RCC with clear-cell or sarcomatoid component and increased recurrence risk post nephrectomy were randomized 1:1 to atezo 1200 mg or pbo IV q3w for 16 cycles or 1 year. A retrospective proteomics analysis was done with an affinity-based proximity extension assay (PEA) panel of ≈3000 analytes to identify circulating proteins with differential abundance patterns in matched serum samples (baseline vs at recurrence). A high sensitivity electrochemiluminescence (ECL) assay was then used to evaluate levels of KIM-1, a membrane glycoprotein overexpressed in clear-cell and papillary RCC, in all available baseline and post-tx serum samples. Outcomes in pts with KIM-1 high (≥86 pg/mL) vs low status at baseline were analyzed. Results: In pts with matched PEA samples (n=73), circulating KIM-1 was identified as the most significantly enriched protein at recurrence vs baseline. Of 778 pts enrolled in IMmotion010, 752 (97%) had baseline KIM-1 data (high: 300 [40%]; low: 452 [60%]). KIM-1–high status was associated with reduced DFS, and pts with KIM-1 high had better DFS with atezo vs pbo (Table). Longitudinal analysis of matched samples showed that in KIM-1–high pts, 9% (12/138) and 26% (36/141) of pts had a ≥30% increase from baseline in KIM-1 levels at Cycle 4 Day 1 with atezo vs pbo, compared with 16% (34/213) and 12% (25/207) in KIM-1-low pts, respectively. A ≥30% KIM-1 increase was associated with worse DFS in both KIM-1–high (atezo HR: 1.68, 95% CI: 0.77, 3.69; pbo HR: 3.53, 95% CI: 2.24, 5.58) and KIM-1–low (atezo HR: 3.56, 95% CI 2.21, 5.75; pbo HR: 3.22, 95% CI: 1.81, 5.70) subgroups. In pts with matched ECL samples (n=103), median KIM-1 levels were higher ( P&lt;0.001) at recurrence (172 pg/mL) than at baseline (79 pg/mL). Conclusions: In IMmotion010, high baseline serum KIM-1 levels were associated with poorer prognosis but improved clinical outcomes with atezo vs pbo. Increased post-tx KIM-1 was associated with worse DFS. These data suggest that circulating KIM-1 may be a non-invasive marker of MRD and disease recurrence and be associated with benefit from atezo in adj RCC. Further investigation of KIM-1 in adj RCC is warranted. Clinical trial information: NCT03024996 . [Table: see text]