Electrochemiluminescence Assay Research Articles

First-in-Human Phase I Trial of a ROR1 Targeting Bispecific T Cell Engager (NVG-111) in Combination with Ibrutinib or As Monotherapy in Subjects with Relapsed Refractory Chronic Lymphocytic Leukaemia (CLL) and Mantle Cell Lymphoma (MCL)

Background: NVG-111 is a first in class, humanized, tandem scFv, ROR1xCD3 bispecific T cell engager that mediates potent killing of ROR1+ tumor by engaging an epitope on the Frizzled domain of ROR1 and redirecting T cell activity via the CD3 binder that has been engineered for lower cytokine release. Methods: This multi-center, open-label, phase I study is evaluating NVG-111 in relapsed/refractory CLL and MCL subjects who have received ≥2 prior systemic therapies. An adaptive trial design was used for dose escalation, starting with accelerated dose titration (ATD) followed by a Bayesian continual reassessment method with overdose control. Each patient received at least one dose of NVG-111, administered as a continuous intravenous infusion (cIV) over 21 days followed by 7 days off drug (=1 cycle). NVG-111 was administered in combination (cohorts 1-4) with ibrutinib to patients who had achieved a stable or partial response to >1 year of ibrutinib therapy, or as monotherapy in patients who had progressed after BTKi / BCL2 (cohort 4b). Measurable residual disease (MRD) was assessed using a flow cytometric method with sensitivity 1e4 (MRD4). Mesoscale discovery electrochemiluminescence assay (MSD-ECLA) was used for pharmacokinetic analysis of NVG-111. Serum cytokine levels were determined using a human high sensitivity cytokine A premixed magnetic Luminex assay. Primary endpoints consisted of safety, tolerability and determination of maximum tolerated dose (MTD)/Recommended Phase 2 Dose (RP2D). Additional end-points included overall response rates as well as duration of response. Results: As of July 2022, 10 subjects (8 males and 2 females, median age 60 years) had been enrolled to the study; three into single subject ATD cohorts, 3 into 30 µg/day flat dosing cohort (cohort 4) and 4 into cohort 4b that entailed a step-up dosing of NVG-111 in cycle 1 only; (first week cycle 1: 3 µg/day, second week, cycle 1: 10 µg/day and week 3, cycle 1: 30 µg/day dose). Eight subjects had CLL and 2 MCL. cIV was well tolerated. Adverse events (AEs) were largely limited to week 1 of cycle 1 and all were reversible. The most common AEs were Grade 1 or 2 nausea (70%), headaches (60%), and fatigue (50%) and thrombocytopenia (30%). Grade 1/2 cytokine release syndrome (CRS) was observed in (40%) of subjects. Grade 3 dose limiting toxicities occurred in two subjects consisting of immune effector cell-associated neurotoxicity syndrome-like symptoms (ICANS) in one (cohort 4) and ALT and AST elevation (cohort 4b) in the other. Both subjects recovered upon stopping NVG-111. Average steady-state serum concentration (Cavg.ss) of NVG-111 in serum of patients who received a 10µg/day and 30µg/day dose were 168-288pg/mL and 425-610pg/mL respectively. This was in line with the predicted drug levels from the single species allometric scaling. Evidence of T cell activation was observed in all evaluable subjects (9/10) with cytokine levels at the 30µg/day dose level being: TNFα, 21±6pg/ml; IL6, 206±178pg/ml; IFNɣ, 7±4pg/ml; and IL10, 47±21pg/ml. Objective clinical responses were observed in 66% of subjects and included 2 MRD4 negative clinical remissions (CR). These two subjects remained in MRD4 negative CRs at 6 months after completion of treatment. Conclusion: Early data shows that NVG-111 is generally well tolerated with a predictable and manageable safety profile. Promising evidence of efficacy was observed which appears to be durable in two subjects with MRD4 negative CR. Dose escalation is ongoing in combination or as monotherapy to determine the MTD/RP2D.

CTIM-13. LOCAL ADOPTIVE CELLULAR IMMUNOTHERAPY WITH CYTOKINE-INDUCED KILLER (CIK) CELLS FOR HIGH GRADE GLIOMAS: A PILOT STUDY WITH LONG-TERM FOLLOW-UP AND POTENTIAL FACTORS FOR SURVIVAL BENEFITS

Abstract BACKGROUND Immunotherapy is emerging as a promising approach for the treatment of high grade gliomas (HGGs). We reported the long-term follow-up of 6 HGG patients treated by local administration of autologous cytokine-induced killer (CIK) cells through Ommaya reservoirs implanted into the tumor cavity following operation. Meanwhile, Gene expression profiles and immune microenvironments of tumors were further compared between the long-term and short -term survivors. METHODS Clinicopathological characteristics and outcomes of patients were reviewed and updated. Gene expression profiles, expressions of cytokines and infiltrations of immune cells in tumors were investigated by RNA sequencing, an electrochemiluminescence assay and immunohistochemistry staining, respectively RESULTS Fever and symptoms of encephaledema occurred in 5 patients after local administration of CIK cells, and could be efficiently relieved by mannitol and dexamethasone. Four patients died from progressive disease during follow-up, and their overall survival ranged from 6 to 26 months. Remarkably, 2 patients have survived more than 200 months without evidence of recurrence. Comparing with the tumors of the short-term survivors, 353 genes which were highly associated with tumor microenvironment immune were differentially expressed (false discovery rate (FDR) < 0.05 and log2 fold change (FC) ≥ 1) in the tumor of the long-term survivor. Higher expressions of cytokines, especially IL-8 and IL-10, were observed in the tumor of the long-term survivor, while the infiltrations of M2 polarized macrophages were significantly higher in the tumors of short-term survivors. CONCLUSION Long-term survival of HGG patients could achieve after local administration of CIK cells into tumor cavity postoperatively. High expressions of cytokines and low infiltrations of M2 polarized macrophages in the tumors potentially benefited the CIK cell therapy.

Cumulative incidence of SARS-CoV-2 infection in the general population of theValencian Community (Spain) after the surge of the Omicron BA.1 variant.

Studies investigating the cumulative incidence of and immune status against SARS-CoV-2 infection provide valuable information for shaping public health decision-making. A cross-sectional study on 935 participants, conducted in the Valencian Community (VC), measuring anti-SARS-CoV-2-receptor binding domain-RBD-total antibodies and anti-Nucleocapsid (N)-IgGs via electrochemiluminescence assays. Quantitation of neutralizing antibodies (NtAb) against ancestral and Omicron BA.1 and BA.2 variants and enumeration of SARS-CoV-2-S specific-IFNγ-producing CD4+ and CD8+ T cells was performed in 100 and 137 participants, respectively. The weighted cumulative incidence was 51.9% (95% confidence interval [CI]: 48.7-55.1) and was inversely related to age. Anti-RBD total antibodies were detected in 97% of participants; vaccinated and SARS-CoV-2-experienced (VAC-ex; n = 442) presented higher levels (p < 0.001) than vaccinated/naïve (VAC-n; n = 472) and nonvaccinated/experienced (UNVAC-ex; n = 63) subjects. Antibody levels correlated inversely with time elapsed since last vaccine dose in VAC-n (Rho, -0.52; p < 0.001) but not in VAC-ex (rho -0.02; p = 0.57). Heterologous booster shots resulted in increased anti-RBD antibody levels compared with homologous schedules in VAC-n, but not in VAC-ex. NtAbs against Omicron BA.1 were detected in 94%, 75%, and 50% of VAC-ex, VAC-n and UNVAC-ex groups, respectively. For Omicron BA.2, the figures were 97%, 84%, and 40%, respectively. SARS-CoV-2-S-reactive IFN-γ T cells were detected in 73%, 75%, and 64% of VAC-ex, VAC-n and UNVAC-ex, respectively. Median frequencies for both T-cell subsets were comparable across groups. In summary, by April 2022, around half of the VC population had been infected with SARS-CoV-2 and, due to extensive vaccination, displayed hybrid immunity.

Non-Myelofibrosis Chronic Myeloproliferative Neoplasm Patients Show Better Seroconversion Rates after SARS-CoV-2 Vaccination Compared to Other Hematologic Diseases: A Multicentric Prospective Study of KroHem.

Disease- and treatment-mediated immunodeficiency might render SARS-CoV-2 vaccines less effective in patients with hematologic diseases. We performed a prospective non-interventional study to evaluate humoral response after one and two doses of mRNA-1273, BNT162b2, or ChAdOx1 nCoV-19 vaccine in 118 patients with different malignant or non-malignant hematologic diseases from three Croatian treatment centers. An electrochemiluminescent assay was used to measure total anti-SARS-CoV-2 S-RBD antibody titers. After one vaccine dose, 20/66 (33%) achieved seropositivity with a median antibody titer of 6.1 U/mL. The response rate (58/90, 64.4%) and median antibody titer (&gt;250 U/mL) were higher after two doses. Seropositivity varied with diagnosis (overall p &lt; 0.001), with the lowest rates in lymphoma (34.6%) and chronic lymphocytic leukemia (52.5%). The overall response rate in chronic myeloproliferative neoplasms (CMPN) was 81.3% but reached 100% in chronic myeloid leukemia and other non-myelofibrosis CMPN. At univariable analysis, age &gt; 67 years, non-Hodgkin's lymphoma, active treatment, and anti-CD20 monoclonal antibody therapy increased the likelihood of no vaccine response, while hematopoietic stem cell recipients were more likely to respond. Age and anti-CD20 monoclonal antibody therapy remained associated with no response in a multivariable model. Patients with the hematologic disease have attenuated responses to SARS-CoV-2 vaccines, and significant variations in different disease subgroups warrant an individualized approach.

Abstract 12349: Can Biomarkers and Risk Scores Be Used to Select Patients With Type 2 Diabetes Mellitus for Testing for Subclinical Left Ventricular Dysfunction

Background: Subclinical LV dysfunction (LVD) identifies heart failure (HF) risk in type 2 diabetes mellitus (T2DM). Hypothesis: Clinical scores (ARIC HF and WATCH-DM) and NTpBNP can be used as screening tests to identify which people should undergo echo. Methods: 803 asymptomatic subjects (69 years [IQR 65-73], 36% female) with T2DM were prospectively recruited from three community-based populations. All underwent a comprehensive echo to identify abnormal global longitudinal strain (GLS &gt; -16%), diastolic function (E/e’ &gt; 14 or e’&lt;8cm/s), left atrial volume index (LAV&gt;34ml/m 2 ) or LV hypertrophy (LV mass index&gt;88g/m 2 (F)&gt;102g/m 2 (M)). ARIC HF risk at 4 years and WATCH-DM scores were calculated from clinical data. NTpBNP was measured using an electro-chemiluminescence assay. Results: In the cohort, 126 (17%) had an abnormal GLS, 16 (2%) elevated E/e’, 266 (33%) had impaired e’, 351 (44%) had increased LAV and 113 (14%) had LV hypertrophy. Median NTpBNP level was 50pg/mL [25-101]. Clinical scores corresponded to expected HF risk (ARIC-HF 8% [4-12]; WATCH-DM 10 points [8-12]). NTpBNP was weakly associated with GLS after adjustments for age, body-mass index, and renal function (β=0.004 [95% CI 0.003 to 0.004], p&lt;0.001). Similar trends were observed with ARIC HF (β=0.07 [0.05 to 0.09], p&lt;0.001) and WATCH-DM (β=0.19 [0.13 to 0.25], p&lt;0.001) in univariable analyses. None of the risk assessment tools provided a satisfactory AUC for detection of abnormal GLS, diastolic indices or LV mass (Table) ( p =0.804). Development of cut-points to optimize sensitivity (to 90%) led to an unacceptably low specificity (Table). Conclusion: Although risk assessment based on clinical scores or biomarkers would be desirable to stratify HF risk in people with T2DM, they show a weak relationship with subclinical LV dysfunction.

Preanalytical stability of SARS-CoV-2 anti-nucleocapsid antibodies.

Anti-nucleocapsid (NC) antibodies are produced in response to SARS-CoV-2 infection. Therefore, they are well suited for the detection of a previous infection. Especially in the case of seroprevalence studies or during the evaluation of a novel in-vitro diagnostic test, samples have been stored at <-70°C (short- and long-term) or 2-10°C (short-term) before analysis. This study aimed to assess the impact of different storage conditions relevant toroutine biobanking on anti-NC antibodies. The preanalytical impact of short-term storage (84 [58-98] days) on <-70°C and for 14days at 2-10°C was evaluated using samples from 111 donors of the MedUni Vienna Biobank. Long-term effects (443 [409-468] days) were assessed using 208 samples from Biobank Graz and 49 samples from Biobank Vienna. Anti-Nucleocapsid antibodies were measured employing electrochemiluminescence assays (Roche Anti-SARS-CoV-2). After short-term storage, the observed changes did not exceed the extent that could be explained by analytical variability. In contrast, results after long-term storage were approximately 20% higher and seemed to increase with storage duration. This effect was independent of the biobank from which the samples were obtained. Accordingly, the sensitivity increased from 92.6 to 95.3% (p=0.008). However, comparisons with data from Anti-Spike protein assays, where these deviations were not apparent, suggest that this deviation could also be explained by the analytical variability of the qualitative Anti-NC assay. Results from anti-NC antibodies are stable during short-term storage at <-70°C and 2-10°C. After long-term storage, a slight increase in sensitivity could notbe ruled out.

A simplified method for monitoring cytokines in wound fluid.

Cytokines in wound fluid are used as surrogates for wound healing in clinical research. The current methods used to collect and process wound fluid are noninvasive but not optimal. The aim of this prospective study was to evaluate a method (NovaSwab) by which wound fluid is collected by a surface swab and eluted in a physiological buffer for subsequent cytokine analysis. Wound fluid from 12 patients with leg ulcers was assessed by NovaSwab at the start (Day 0) and at the end of a 23-h collection period of wound fluid retained by foam oblates beneath an occlusive film dressing (Day 1). GM-CSF, IL-1α, IL-1β, IL-6, IL-8, PDGF-AA, TNF-α and VEGF levels were measured by multiplex and electrochemiluminescence assays. IL-1α (2.4×), IL-1β (2.0×) and IL-8 (1.8×) levels increased from Day 0 to Day 1 as detected by NovaSwab, indicating local production of these polypeptides in the wounds. On Day 1, the NovaSwab method yielded higher levels of IL-1α (4.0×), IL-1β (2.7×) and IL-6 (2.7×), and 35% lower levels of VEGF than those in wound fluid accumulated for 23 h in foam oblates (on average, 5ml of wound fluid). In vitro experiments showed that the investigated cytokines in cell-free wound fluid were recovered in a quantitative manner by the NovaSwab method. We conclude that the method presented here is a promising research tool to study the kinetics of soluble cytokines over the course of wound healing. More studies are needed to determine the interobserver variation and reproducibility of the NovaSwab method.

Reusable molecularly imprinted electrochemiluminescence assay for kanamycin based on ordered mesoporous carbon loaded with indium oxide nanoparticles and carbon quantum dots.

Ahighly sensitive kanamycin electrochemiluminescence (ECL) switch sensor was constructed. A signal element consisting of ordered mesoporous carbon loaded with indium oxide nanoparticles/carbon quantum dots (OMC/In2O3/C-dots) was assembled on the surface of a gold electrode. Then, a molecularly imprinted polymer (MIP) was prepared on the modified electrode surface using kanamycin as the template molecule and o-aminophenol as the functional monomer. After kanamycin elution, the prepared sensor retained specific kanamycin recognition sites. OMC/In2O3 effectively amplified the ECL signal of the C-dots, thereby enhancing the detection sensitivity, whereas kanamycin quenched the signal. Therefore, the imprinted sites acted as a switch, providing a new method for detecting kanamycin. Under the optimal experimental conditions, the concentration of kanamycin was proportional to the degree of ECL quenching within a linear range of 5-4500 × 10-12mol L-1 at 0.8V (vs. Ag/AgCl electrode electrode), and the detection limit was 5.8 × 10-13mol L-1. Whenapplied to the detection of kanamycin in actual samples, such as chicken, duck, pork, and milk, the recovery for spiked samples was in the range92.7-110%.

16 SARS-COV-2 INFECTION AND VACCINATION PATTERNS DETERMINE LONG-TERM ANTIBODY RESPONSES IN NURSING HOME RESIDENTS: DATA FROM NH-COVAIR

Abstract Background Older Nursing Home Residents (NHRs) are at greatest risk of morbidity and mortality from SARS-CoV-2, particularly in the context of both waning vaccine efficacy and the emergence of Variants-of-Concern (VOCs). However, the determinants of long-term vaccine-induced protective antibody responses are yet to be determined in this group. Methods NH-COVAIR recruited older NHRs for comprehensive clinical and frailty (NH-FRAIL) assessment. Blood samples were obtained pre-vaccination, at 6-weeks and 6-months following primary vaccination and 6-months following booster vaccination. Antibody titres were measured using both an electrochemiluminescence assay and a custom bead-based array (Luminex™) to measure antibody titre and avidity for Wuhan strain/major VOC antigens. Stepwise adjusted linear regression (log-transformed) assessed longitudinal determinants of vaccine-induced antibody responses. Results Of 86 participants (81.1 ± 10.8 years; 65% female), just under half (45.4%) had evidence of previous SARS-CoV-2 infection. All NHRs mounted a significant antibody-response to vaccination at 5 weeks followed by a significant decrease in antibody titre by 6 months. Previous SARS-CoV-2 infection was the strongest predictor of antibody waning at all timepoints (β: 3.59; 2.89, 4.28; P &amp;lt; 0.001 for 6-months). Independent of infection history, both age (β: –0.05; –0.08, –0.02; p&amp;lt;0.001) and frailty (β: –0.22; –0.33, –0.11; p&amp;lt;0.001) were associated with faster antibody waning at 6-months. Cross-reactivity and avidity were significantly lower for Beta (B.1.351) and Gamma (P.1) VOC strains (all p&amp;lt;0.001). Additionally, there was faster antibody waning and significantly reduced antibody avidity to Beta and Gamma VOCs in SARS-CoV-2 naïve NHRs. Conclusion Older NHRs are capable of mounting protective antibody responses to SARS-CoV-2 vaccination. Responses were more durable, with a greater cross-reactivity to and avidity for VOCs in those with previous SARS-CoV-2 infection. Increasing age and greater frailty in NHRs was associated with faster antibody waning. Our findings support ongoing serological surveillance and use of additional vaccine doses in older NHRs, particularly in those without previous SARS-CoV-2 exposure.

Topical review- salivary biomarkers in chronic muscle pain.

Muscle related temporomandibular disorders (myogenous TMD), one of the most common orofacial pain conditions, is characterized by facial pain and often accompanied by jaw movement limitations. Although the underlying biological mechanisms are still unclear, a cluster of proteins and peptides is assumed tobe involved in the pathophysiology. These proteins andpeptides may be measured in a simple non-invasive saliva sample. This work investigated whether saliva can be used to sample algogenic substances that can serve asmolecular biomarkers for TMD myalgia. Saliva and blood samples were collected from healthy individuals (n=69) and patients diagnosed with TMD myalgia (n=39) according to the Diagnostic Criteria for TMD. Unstimulated and stimulated whole, parotid, and sublingual saliva were analysed. The protein profiles were investigated using two-dimensional gel electrophoresis followed by identification with liquid chromatography tandem mass spectrometry. Levels of nerve growth factor (NGF), calcitonin gene-related peptide (CGRP), and brain derived neuro-tropic factor (BDNF) were determined using western blotting based technology and multiplex electro-chemiluminescence assay panel. Glutamate, serotonin, and substance p (SP) were determined using commercially available methods. Different saliva collection approaches resulted insignificant differences in the protein profile as well as inthe expression of NGF, BDNF, CGRP, SP, and glutamate. Stimulated whole saliva showed least variability in protein concentration (35%) and was correlated to plasma levels ofglutamate. Unlike SP and glutamate, NGF and BDNF expressed a rhythmic variation in salivary expression withhigher levels in the morning (p<0.05). Patients with a diagnosis of TMD myalgia had significantly higher levels of salivary glutamate but lower salivary NGF and BDNF compared to controls; in addition, the lower NGF and BDNF levels correlated to psychological dysfunction. Thequantitative proteomics data revealed 20 proteins that were significantly altered in patients compared to controls. The identified proteins are involved in metabolic processes, immune response, and stress response. Dissimilarities in protein profile and clinical variables were observed between TMD myalgia and myofascial pain. The work highlights the importance of consistency in saliva collection approaches, including thetiming of the collection. It displayed significant changes in pain specific mediators and protein profile in TMD myalgia and furthermore dissimilarities between subclasses indicating different pathophysiology. After extensive validation, potential salivary biomarkers can becombined with clinical features to better understand and diagnose TMD myalgia.

Erythrocytic alpha-synuclein in early Parkinson's disease: A 3-year longitudinal study

BackgroundEarly diagnosis of Parkinson's disease (PD) could significantly improve outcomes for patients and future disease-modifying treatments. Several studies have revealed that α-synuclein levels in peripheral erythrocytes are associated with PD, but the diagnostic value in early PD is still unknown. MethodsThis study included both cross-sectional and longitudinal design. The subjects included 45 patients with early PD and 79 age-matched healthy controls. Participants were re-examined with repeated blood collection and clinical assessments after 3 years. The electrochemiluminescence assay was used to measure total and oligomeric α-synuclein levels respectively. The diagnostic value of erythrocytic α-synuclein for early PD was determined by receiver operator characteristic (ROC) curve. Correlations between RBC α-synuclein levels and changes over 3 years in clinical characteristic scores were further investigated with a linear regression. ResultsTotal and oligomeric α-synuclein levels in erythrocyte were significantly increased in early PD groups compared with control group (Total α-synuclein, p < 0.001; Oligomer, p < 0.001). Levels of total and oligomeric α-synuclein in erythrocytes were correlated with MDS-UPDRS III scores in early PD (Total α-synuclein, p = 0.008; Oligomer, p = 0.037). After adjusting for age, gender and dopaminergic medication, an association was found between higher erythrocytic oligomeric α-synuclein levels at baseline and greater increase in MDS-UPDRS III scores over 3 years (p = 0.007). ConclusionOur study suggests that total and oligomeric α-synuclein in erythrocyte were elevated even in the initial motor stage of PD. Higher erythrocytic oligomeric α-synuclein levels at baseline predicts a faster clinical decline over time in patients with early PD.

Relationship between cerebrospinal fluid biomarkers of inflammation and tissue damage in primary progressive multiple sclerosis

Background and ObjectivesIt is unclear to what extent intrathecal inflammation contributes to the pathogenesis in primary progressive multiple sclerosis (PPMS). We conducted an exploratory study to investigate the degree of intrathecal inflammation and its association with biomarkers of disease activity and severity in patients with PPMS. MethodsWe included patients with PPMS who participated in a randomized controlled trial conducted at the Danish Multiple Sclerosis Center, patients with relapsing-remitting multiple sclerosis (RRMS) and healthy controls. We analyzed concentrations of a panel of cytokines in CSF using electrochemiluminescence assays. We then explored the relationship between cytokines found in increased CSF concentrations in patients with PPMS (compared with healthy controls) with CSF concentrations of neurofilament light chain (NFL) and myelin basic protein (MBP), IgG-index, and magnetic resonance imaging (MRI) metrics (volume, magnetization transfer ratio and diffusion tensor imaging) from lesions, normal-appearing white matter, and cortical grey matter. ResultsWe included 59 patients with PPMS, 40 patients with RRMS, and 21 healthy controls. In patients with PPMS, CSF concentrations of CC chemokine ligand 3 (CCL-3), CXC chemokine ligand 8 (CXCL-8), CXCL-10, interleukin (IL)-10, IL-15, and vascular endothelial growth factor (VEGF)-A were increased compared with healthy controls and comparable with CSF concentrations in patients with RRMS. In addition, patients with PPMS had increased CSF concentrations of IL-12p40, IL-17A, tumor necrosis factor (TNF)-α, and lymphotoxin (LT)-α compared with healthy controls, but concentrations of these cytokines were even higher in patients with RRMS. For the remaining seven cytokines (CCL22, interferon-γ, IL-5, IL-7, IL-16, IL-22, IL-27), we found no difference between patients with PPMS and healthy controls. CSF concentrations of NFL and MBP correlated weakly with concentrations of IL-15, while the remaining proinflammatory cytokines were not associated with CSF concentrations of NFL or MBP. The IgG-index correlated with four cytokines (IL-10, IL-12p40, TNF-α, and LT-α). We did not observe any significant associations between MRI metrics and CSF biomarkers of inflammation. DiscussionIn this exploratory study, we found few and weak associations between intrathecal inflammation and the extent of neuroaxonal damage and demyelination, and no associations between intrathecal inflammation and MRI metrics, in patients with PPMS. Our findings suggest that, for patients with PPMS, these measures of intrathecal inflammation are not associated with the extent of neuroaxonal injury, demyelination, and disease severity, and these processes may therefore have less relevance in PPMS than in relapsing forms of MS.

Electrochemiluminescence in paired signal electrode (ECLipse) enables modular and scalable biosensing.

Electrochemiluminescence (ECL) has an inherently low background and enables precise chemical reactions through electrical control. Here, we report an advanced ECL system, termed ECLipse (ECL in paired signal electrode). We physically separated ECL generation from target detection: These two processes were carried out in isolated chambers and coupled through an electrode. The strategy allowed us to minimize cross-chemical reactions, design electrodes for high ECL signals, and integrate multiple sensors in a chip. As a proof of concept, we implemented an eight-plex ECLipse and applied it to detect host factors in human plasma. ECLipse achieved higher signal-to-noise ratio than conventional ECL assays and was >7000-fold more sensitive than enzyme-linked immunosorbent assay. In a pilot clinical study, we could detect septic conditions by measuring host factors [i.e., interleukin-3 (IL-3), IL-6, and procalcitonin (PCT)]. ECLipse assay further revealed distinct IL-3 and IL-6 patterns in patients with severe acute respiratory syndrome coronavirus 2 infection.

Vaccine-Induced T-Cell and Antibody Responses at 12 Months after Full Vaccination Differ with Respect to Omicron Recognition.

More than a year after the first vaccines against the novel SARS-CoV-2 were approved, many questions still remain about the long-term protection conferred by each vaccine. How long the effect lasts, how effective it is against variants of concern and whether further vaccinations will confer additional benefits remain part of current and future research. For this purpose, we examined 182 health care employees—some of them with previous SARS-CoV-2 infection—12 months after different primary immunizations. To assess antibody responses, we performed an electrochemiluminescence assay (ECLIA) to determine anti-spike IgGs, followed by a surrogate virus neutralization assay against Wuhan-Hu-1 and B.1.1.529/BA.1 (Omicron). T cell response against wild-type and the Omicron variants of concern were assessed via interferon-gamma ELISpot assays and T-cell surface and intracellular cytokine staining. In summary, our results show that after the third vaccination with an mRNA vaccine, differences in antibody quantity and functionality observed after different primary immunizations were equalized. As for the T cell response, we were able to demonstrate a memory function for CD4+ and CD8+ T cells alike. Importantly, both T and antibody responses against wild-type and omicron differed significantly; however, antibody and T cell responses did not correlate with each other and, thus, may contribute differentially to immunity.

Dual-wavelength electrochemiluminescence biosensor based on a multifunctional Zr MOFs@PEI@AuAg nanocomposite with intramolecular self-enhancing effect for simultaneous detection of dual microRNAs

Rapid parallel detection of multi-targets has always been an exploration aim in electrochemiluminescence (ECL) assays. Herein, a multifunctional nanocomposite of Zr metal-organic frameworks (MOFs) @PEI@AuAg nanoclusters (NCs) with intense and stable dual-wavelength ECL was synthesized for the first time, and used to construct a new ECL biosensor for rapid simultaneous detection of dual targets. Notably, the novel ECL emitter Zr MOFs with high-performance was not only integrated with a co-reactant polyethyleneimine (PEI) to form a unique intramolecular self-enhancing structure, but also loaded a large number of another ECL emitter AuAg NCs, furthermore, AuAg NCs with superior electron transfer property can much enhance the electrical conductivity of the composites, thus achieving the goal of “killing three birds with one stone”. Moreover, a unique stable and rigid three-dimensional DNA tetrahedron (TDN) structure was connected with two quenching probes BHQ1 and BHQ3 and immobilized on the composites-modified electrode, so ECL emission of the nanocomposites at two wavelengths of 535 nm and 644 nm were both quenched by resonance energy transfer (RET). In the presence of target miRNAs, the efficient DNA cycling double-amplification processes were performed by using exonuclease (T7 Exo) combined with DNA Walker, thus both quenching groups were separated to restore the ECL at two wavelengths, achieving simultaneous and rapid ECL detection of two miRNAs. Therefore, this present work not only opens a unique nanocomplex with dual wavelength ECL and self-enhancing performance, but also develops a highly sensitive ECL biosensor with promising value for rapid multi-target analysis in clinical fields.

Circulating CD45+EpCAM+ cells as a diagnostic marker for early-stage primary lung cancer.

Lung cancer is a highly prevalent type of cancer, accounting for 11.6% of all cancer incidences. Early detection and treatment can significantly improve the survival rate and quality of life of patients; however, there is no accurate, effective, and easy-to-use test for early lung cancer screening. In this study, flow cytometry was used to detect the presence of CD45+EpCAM+ cells in tumor tissues and peripheral blood mononuclear cells (PBMCs) in patients with lung cancer. Moreover, the proportion of CD45+EpCAM+ cells in PBMCs of patients with lung cancer was found to be significantly higher than that of healthy volunteers. Tumor-related serum markers level was also measured in the peripheral blood of these patients using an electrochemiluminescence assay. The correlation between CD45+EpCAM+ cells, carcinoembryonic antigen (CEA), and lung cancer was investigated using receiver operating characteristic (ROC) curve analysis, which showed the sensitivity and specificity of the CD45+EpCAM+ cell to be 81.58% and 88.89%, respectively. Further analysis yielded an area under the ROC curve (ROC/area under the curve [AUC]) of 0.845 in patients PBMCs with lung cancer, which was slightly higher than that of CEA (0.732). Therefore, the detection of CD45+EpCAM+ cells in PBMCs may be helpful for the early screening and auxiliary diagnosis of lung cancer.

Is serum NSE a useful biomarker in multiple system atrophy?

Objective: Multiple system atrophy (MSA) is a rapidly progressive and devastating neurodegenerative disease with limited lifespan. Biomarkers in diagnosing and evaluating the prognosis of MSA is a rapidly developing field. Neuron-specific enolase (NSE) is a biomarker of neuronal damage applicable in many neurological disorders. This study aimed to determine the usefulness of serum NSE as biomarker of MSA. Methods: In this case-control study involving 50 patients with MSA, electrochemiluminescence assay was used to investigate whether serum NSE levels. The disease severity of patients was assessed by Unified System Atrophy Rating Scale (UMSARS part II). The correlation between NSE levels and clinical features in MSA patients was analyzed. Results: The NSE levels were higher in MSA patients (11.59±2.52 ng/mL) than the control group (11.30±2.73 ng/mL); however, the difference was not statistically significant (P=0.581). In addition, no statistically significant difference was detected in the NSE levels between the parkinsonian (MSA-P) and cerebellar subtypes (MSA-C). No distinct difference was also detected between males and females of MSA patients. Furthermore, no correlations were established between NSE levels and age, disease duration, and UMSARS II score. Conclusions: This study suggest that serum NSE is not a useful biomarker of MSA. Future studies should examine whether there is a role for CSF NSE in reflecting neuronal damage in MSA patients.

Electrochemiluminescent (ECL) biosensor for Burkholderia pseudomallei based on cobalt-doped MOF decorated with gold nanoparticles and N-(4-aminobutyl)-N-(ethylisoluminol).

A multifunctional catalytic nanomaterial (Co-MOF@AuNP@ABEI) composed of cobalt-doped metal-organic frameworks (Co-MOF), gold nanoparticles (AuNP), and N-(4-aminobutyl)-N-(ethylisoluminol) (ABEI) is reported. Co-MOF@AuNP@ABEI exhibits high synergistic and zero-distance catalytic properties, which are beneficial to the improvement of the detection sensitivity of an electrochemiluminescent (ECL) biosensor. After coupling with the ECL system and 3D magnetic walking nanomachine amplification strategy, the Co-MOF@AuNP@ABEI can achieve an ultrasensitive ECL assay of Burkholderia pseudomallei with the limit of detection (LOD) of 60.3 aM, which is 2 and 4 orders of magnitude lower than individual ECL system without the nanomachine (4.97 fM) and individual walking nanomachine (340 fM), and superior to the pathogenic bacteria analyses in the previous report. Moreover, the LOD of the proposed ECL detection system for the determination of B. pseudomallei in serum sample was as low as 9.0CFUmL-1. The relative standard deviations (RSD) of ECL intensity for the detection of five B. pseudomallei-spiked serum samples were 4.02%, 0.84%, 0.84%, 1.55%, and 0.21%, respectively. The recoveries of the ECL biosensor for the detection of B. pseudomallei DNA-spiked serum samples were 93.63 ~ 107.83%. Therefore, this work demonstrated that the developed multifunctional catalytic nanomaterial with synergistic and zero-distance catalytic properties can be used as excellent ECL signal reporter to improve the detection sensitivity of ECL biosensor.

Serum IL-13 Predicts Response to Golimumab in Bio-Naïve Ulcerative Colitis.

A certain number of patients with ulcerative colitis (UC) are refractory to anti-TNF-α antibodies; biomarkers are thus needed to predict treatment efficacy. This study aimed to evaluate whether serum biomarkers that were reported to be associated with UC or anti-TNF-α antibody could predict the response to golimumab, a human anti-TNF-α monoclonal antibody, in bio-naïve patients with UC. We prospectively enrolled 23 consecutive patients with UC who were treated with golimumab. Serum samples were collected before the first golimumab dose. Eleven molecules were measured by electrochemiluminescence (ECL) or enzyme-linked immunosorbent assay (ELISA) and their association with efficacy after 10 weeks of golimumab treatment. Among the serum biomarkers, IL-13 levels were significantly higher in the non-remission group than in the remission group (p = 0.014). IL-15 levels were significantly lower in the non-response group than in the response group (p = 0.04). For clinical remission at week 10, the IL-13 0.20 concentration of pg/mL was associated with a sensitivity and specificity of 82.4% and 83.3%, respectively. Serum IL-13 may be a biomarker to predict golimumab efficacy in biologic-naïve patients with UC, and thus may help to tailor personalized treatment strategies.

Correlation between Results of Semi-Quantitative and Quantitative Tests for Hepatitis B Virus Surface Antigen among Patients Achieving Viral Suppression with Antiviral Treatment.

Background: Hepatitis B virus (HBV) infection remains a threat to global public health. Serum hepatitis B surface antigen (HBsAg) has been used in screening for HBV infection. Quantitative HBsAg assays are useful for monitoring the natural history of HBV infection and its response to therapy. The aim of this study was to determine the relationship between quantitative (qHBsAg; IU/mL) and semi-quantitative (sqHBsAg; signal-to-cutoff ratio [S/Co]) HBsAg titers in patients with chronic hepatitis B (CHB). Methods: We retrospectively included 284 samples with HBV DNA < 20 IU/mL from patients who had simultaneous qHBsAg (using electrochemiluminescence assay) and sqHBsAg tests. Patients were grouped according to their serum HBV-envelope antigen (HBeAg) status (HBeAg-negative, n = 239 and HBeAg-positive, n = 45). The Spearman test was used to analyze the correlation between the quantitative and semi-quantitative assays. Results: There was a significant linear correlation between sqHBsAg and qHBsAg in the HBeAg-negative patients (qHBsAg [IU/mL] = 0.0094 × sqHBsAg [S/Co]1.323; adjusted R2 = 0.8445; p < 0.001). There was a substantial hook effect in the assays from the HBeAg-positive patients, so we performed a stratified analysis according to qHBsAg <1000 IU/mL or ≥1000 IU/mL and found a significant positive linear correlation between sqHBsAg S/Co and qHBsAg (qHBsAg [IU/mL] = 0.072 × sqHBsAg [S/Co]1.331; adjusted R2 = 0.7878; p < 0.001) in HBeAg-positive patients with qHBsAg titers of <1000 IU/mL and a significant negative correlation in HBeAg-positive patients with qHBsAg titers of ≥1000 IU/mL (qHBsAg [IU/mL] = 8.987 × 1014 × sqHBsAg [S/Co]−3.175; adjusted R2 = 0.6350; p < 0.001). Conclusions: There was a highly linear, positive correlation between qHBsAg and sqHBsAg in HBeAg-negative CHB patients. The hook effect led to a negative correlation in HBeAg-positive CHB patients with qHBsAg titers ≥1000 IU/mL.