scholarly journals 645. Singulex Clarity Norovirus Assay (In Development) Provides Ultrasensitive Detection of Norovirus Genogroups I and II

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S297-S298
Author(s):  
Gipshu Dave ◽  
Phoebe Katzenbach ◽  
Johanna Sandlund ◽  
Joel Estis ◽  
Ali Mukherjee ◽  
...  
Keyword(s):  
Single Molecule ◽  
Operating Characteristic ◽  
High Sensitivity ◽  
Excellent Performance ◽  
C Statistic ◽  
Stool Samples ◽  
Capture Antibody ◽  
Automated Platform ◽  
Paramagnetic Microparticles ◽  
Detection Reagent

Abstract Background Commercially available enzyme immunoassays (EIAs) for detection of norovirus antigen have poor sensitivity and are limited to use in investigations of a gastroenteritis outbreak. Hence, there remains a need for a standalone high-sensitivity assay that enables rapid and accurate detection of norovirus antigen. Methods The Singulex Clarity norovirus assay is currently in development for use on the Singulex Clarity® system (Singulex Inc., Alameda, CA, USA), a fully-automated platform powered by Single Molecule Counting technology (registered with the FDA and CE marked). The assay uses paramagnetic microparticles bound to capture antibody and a fluorescently labeled reporter antibody to detect virion capsid protein of norovirus genogroups I (GI) and II (GII) in the stool. For the development of Clarity Norovirus assay, diagnostic performance of 4 antibody pairs (as Capture and Detection reagent) were evaluated by testing 137 stool samples from patients with suspected norovirus infection. Samples were sourced from three providers: (1) 90 genotyped samples of which 75 were positive (19 different genotypes) and 15 were negative by the CDC assay, (2) 3 samples positive and 5 samples negative by the BioFire® FilmArray® Gastrointestinal Panel, and (3) 39 samples negative by a lab-developed test using Cepheid reagents (SmartCycler®). Results From all the antibody pairs tested, one of the pairs had best performance with the area under the receiver operating characteristic (AuROC) curve demonstrating a C-Statistic of 0.959 (95% CI 0.921–0.997), compared with AuROC C-statistic of 0.943 (95% CI 0.896–0.990), 0.871 (95% CI 0.807–0.936), and 0.914 (95% CI 0.863–0.964) for the three other pairs. The Clarity assay detected all 19 different genotypes tested (figures). Conclusion The ultrasensitive and rapid Clarity norovirus assay (in development) for detection of GI and GII demonstrated excellent performance with one of the antibody pairs tested and detected all 19 tested genotypes. The Clarity assay may offer a standalone solution for norovirus diagnostics. Disclosures All authors: No reported disclosures.

scholarly journals 2357. Toxin Detection Using Single Molecule Counting Technology: The Best of Both Worlds?

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S812-S812
Author(s):  
Michael Perry ◽  
Lee Graham ◽  
Sweta Parida ◽  
Phoebe Katzenbach ◽  
Jose Luis Baptista Baeza ◽  
...  
Keyword(s):  
Single Molecule ◽  
Neutralization Assay ◽  
Accurate Diagnosis ◽  
Excellent Performance ◽  
Toxin Concentration ◽  
Toxin B ◽  
Clinical Sensitivity ◽  
Percent Agreement ◽  
Stool Samples ◽  
Membrane Type

Abstract Background Accurate diagnosis of CDI remains challenging as there is no standalone laboratory test with adequate clinical sensitivity and specificity. Thus, many clinical laboratories currently employ a multistep algorithm incorporating a sensitive screening test followed by a specific toxin test. An automated ultrasensitive toxin immunoassay (Singulex Clarity® C. difficile toxins A/B assay) has demonstrated excellent performance compared with cell cytotoxicity neutralization assay (CCNA). In this study, the Clarity assay was evaluated relative to glutamate dehydrogenase (GDH), toxin EIA, toxin B gene PCR, multistep algorithms, and C. difficile culture with ribotyping. Methods Residual clinical stool samples (n = 293) were collected from patients with suspected CDI. The samples were tested on-site with GDH (C. DIFF CHEK™-60), PCR (EntericBio realtime® C. difficile assay), a membrane-type toxin EIA (Tox A/B Quik Chek®), and culture and ribotyping. In total, 188 samples were tested with GDH and 239 samples were tested by PCR. All PCR-positive samples (n = 148) and prospectively tested GDH samples (n = 97) were tested with the toxin EIA. Culture and ribotyping information were available for 205 samples. Results Three of the samples tested gave no result using the Clarity assay and were excluded from the analysis. The Singulex Clarity C. difficile toxins A/B assay had high positive percent agreement (PPA) and low negative percent agreement (NPA) compared with toxin EIA and multistep algorithms ending with toxin EIA. The Clarity assay had high NPA and low PPA compared with PCR, GDH, and the multistep algorithm ending with PCR (figure). Less than 70% of the detected C. difficile PCR positive samples had toxins present. There was no difference in toxin concentration between the ribotypes. Conclusion The Clarity assay had strong PPA compared with toxin EIA and strong NPA compared with PCR. The low NPA and PPA compared with toxin EIA and PCR, respectively, may reflect the poor sensitivity of current toxin EIAs and low specificity of PCR. The Clarity assay detected 30 different ribotype strains, and less than 70% of samples (by PCR) or strains (by ribotyping) had toxins present. The Clarity assay may be considered for use as a standalone test for CDI diagnosis. Disclosures All authors: No reported disclosures.

scholarly journals Generation and characterization of monoclonal antibodies against mature hepcidin and its application to neutralization and quantitative alteration assay

2021 ◽  
Vol 85 (2) ◽  
pp. 340-350
Author(s):  
Shinji Sakamoto ◽  
Mika Kirinashizawa ◽  
Yumi Mohara ◽  
Yoshihiro Watanabe
Keyword(s):  
Monoclonal Antibodies ◽  
Sandwich Elisa ◽  
High Sensitivity ◽  
Cellular Membrane ◽  
Average Concentration ◽  
Cellular Iron ◽  
Capture Antibody ◽  
Quantitative Alteration ◽  
Detection Reagent

ABSTRACT Hepcidin regulates the quantity of ferroportin (FPN) on cellular membrane. In our cell assay expressing ferroportin labeled with green fluorescence, FPN was internalized and degraded only after treatment with hepcidin-25, not hepcidin-22 or hepcidin-20, leading to accumulation of cellular iron. Thus we generated murine monoclonal antibodies (mAbs) against hepcidin-25, and then characterized and validated their functions. Among them, several mAbs showed a neutralizing activity that may prevent ferroportin internalization induced by hepcidin-25. To measure hepcidin level in various fluids, mAbs specific for human and rat hepcidin-25 were selected. As for rat, a sandwich ELISA developed using clone rHN1 as capture antibody and biotinylated clone mHW1 as a detection reagent had high sensitivity, allowing for the detection of 1-100 ng/mL of hepcidin-25. Rat hepcidin-25 level in plasma was measured at an average concentration of 63.0 ng/mL in healthy condition, and at 218.2 ng/mL after stimulation of lipopolysaccharide.

Direct Read and Quantify Damage Nucleotide from an Oligonucleotide Using a Single Molecule Interface

2019 ◽  
Author(s):  
Jiajun Wang ◽  
Meng-Yin Li ◽  
Jie Yang ◽  
Ya-Qian Wang ◽  
Xue-Yuan Wu ◽  
...  
Keyword(s):  
Dna Sequencing ◽  
Single Molecule ◽  
Genetic Diseases ◽  
High Sensitivity ◽  
Dna Lesions ◽  
Lesion Detection ◽  
The Novel ◽  
Structural Variations ◽  
Dna Lesion ◽  
Base Lesion

DNA lesion such as metholcytosine(<sup>m</sup>C), 8-OXO-guanine(<sup>O</sup>G), inosine(I) <i>etc</i> could cause the genetic diseases. Identification of the varieties of lesion bases are usually beyond the capability of conventional DNA sequencing which is mainly designed to discriminate four bases only. Therefore, lesion detection remain challenge due to the massive varieties and less distinguishable readouts for minor structural variations. Moreover, standard amplification and labelling hardly works in DNA lesions detection. Herein, we designed a single molecule interface from the mutant K238Q Aerolysin, whose confined sensing region shows the high compatible to capture and then directly convert each base lesion into distinguishable current readouts. Compared with previous single molecule sensing interface, the resolution of the K238Q Aerolysin nanopore is enhanced by 2-order. The novel K238Q could direct discriminate at least 3 types (<sup>m</sup>C, <sup>O</sup>G, I) lesions without lableing and quantify modification sites under mixed hetero-composition condition of oligonucleotide. Such nanopore could be further applied to diagnose genetic diseases at high sensitivity.

scholarly journals Relationship between serum angiopoietin-like 4 levels and recurrence of atrial fibrillation

2021 ◽  
Vol 49 (2) ◽  
pp. 030006052098839
Author(s):  
Zhongping Ning ◽  
Xinming Li ◽  
Xi Zhu ◽  
Jun Luo ◽  
Yingbiao Wu
Keyword(s):  
Atrial Fibrillation ◽  
Catheter Ablation ◽  
Operating Characteristic ◽  
Characteristic Curve ◽  
High Sensitivity ◽  
Peroxisome Proliferator ◽  
C Reactive Protein ◽  
Peroxisome Proliferator Activated Receptor ◽  
Reactive Protein ◽  
New Onset

Objective To investigate the association between serum angiopoietin-like 4 (ANGPTL4) levels and recurrence of atrial fibrillation (AF) after catheter ablation. Methods This retrospective study recruited patients with AF undergoing catheter ablation and they were divided into two groups (new-onset AF group and recurrent AF group). Demographic, clinical, and laboratory parameters were collected. Results A total of 192 patients with AF were included, including 69 patients with recurrence of AF. Serum ANGPTL4 levels were lower in patients with recurrent AF than in those with new-onset AF. Serum ANGPTL4 levels were positively correlated with superoxide dismutase and peroxisome proliferator-activated receptor γ, and negatively correlated with the CHA2DS2-VASC score, left atrial diameter, and levels of brain natriuretic peptide, malondialdehyde, high-sensitivity C-reactive protein, and interleukin-6. The receiver operating characteristic curve showed that the best cut-off for recurrent AF was serum ANGPTL4 levels  < 19.735 ng/mL, with a sensitivity and specificity of 63.9% and 74.5%, respectively. Serum ANGPTL4 levels were significantly associated with recurrence and new onset of AF (odds ratio, 2.241; 95% confidence interval, 1.081–4.648). Conclusions Serum ANGPTL4 levels are lower in patients with recurrent AF than in those with new-onset AF, and are associated with cardiac hypertrophy, oxidative stress, and inflammation.

scholarly journals Nanopore Technology for the Application of Protein Detection

Nanomaterials ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 1942
Author(s):  
Xiaoqing Zeng ◽  
Yang Xiang ◽  
Qianshan Liu ◽  
Liang Wang ◽  
Qianyun Ma ◽  
...  
Keyword(s):  
Single Molecule ◽  
Protein Detection ◽  
High Sensitivity ◽  
Single Molecule Detection ◽  
Sequence Detection ◽  
Fast Detection ◽  
Material Basis ◽  
Sensing Technology ◽  
Detection Technology ◽  
Structure Recognition

Protein is an important component of all the cells and tissues of the human body and is the material basis of life. Its content, sequence, and spatial structure have a great impact on proteomics and human biology. It can reflect the important information of normal or pathophysiological processes and promote the development of new diagnoses and treatment methods. However, the current techniques of proteomics for protein analysis are limited by chemical modifications, large sample sizes, or cumbersome operations. Solving this problem requires overcoming huge challenges. Nanopore single molecule detection technology overcomes this shortcoming. As a new sensing technology, it has the advantages of no labeling, high sensitivity, fast detection speed, real-time monitoring, and simple operation. It is widely used in gene sequencing, detection of peptides and proteins, markers and microorganisms, and other biomolecules and metal ions. Therefore, based on the advantages of novel nanopore single-molecule detection technology, its application to protein sequence detection and structure recognition has also been proposed and developed. In this paper, the application of nanopore single-molecule detection technology in protein detection in recent years is reviewed, and its development prospect is investigated.

scholarly journals Single-copy detection of somatic variants from solid and liquid biopsy

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ana-Luisa Silva ◽  
Paulina Klaudyna Powalowska ◽  
Magdalena Stolarek ◽  
Eleanor Ruth Gray ◽  
Rebecca Natalie Palmer ◽  
...  
Keyword(s):  
Real Time ◽  
Single Molecule ◽  
Real Time Pcr ◽  
Clinical Decision Making ◽  
High Sensitivity ◽  
Clinical Decision ◽  
Single Copy ◽  
Turnaround Time ◽  
Copy Detection ◽  
Allele Specific

AbstractAccurate detection of somatic variants, against a background of wild-type molecules, is essential for clinical decision making in oncology. Existing approaches, such as allele-specific real-time PCR, are typically limited to a single target gene and lack sensitivity. Alternatively, next-generation sequencing methods suffer from slow turnaround time, high costs, and are complex to implement, typically limiting them to single-site use. Here, we report a method, which we term Allele-Specific PYrophosphorolysis Reaction (ASPYRE), for high sensitivity detection of panels of somatic variants. ASPYRE has a simple workflow and is compatible with standard molecular biology reagents and real-time PCR instruments. We show that ASPYRE has single molecule sensitivity and is tolerant of DNA extracted from plasma and formalin fixed paraffin embedded (FFPE) samples. We also demonstrate two multiplex panels, including one for detection of 47 EGFR variants. ASPYRE presents an effective and accessible method that simplifies highly sensitive and multiplexed detection of somatic variants.

scholarly journals Dysbiosis characteristics of gut microbiota in cerebral infarction patients

2020 ◽  
Vol 11 (1) ◽  
pp. 124-133
Author(s):  
Hao Li ◽  
Xiaohui Zhang ◽  
Dengdeng Pan ◽  
Yongqiang Liu ◽  
Xuebing Yan ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Cerebral Infarction ◽  
Operating Characteristic ◽  
National Institutes Of Health ◽  
Diagnostic Model ◽  
Healthy Controls ◽  
Characteristic Analysis ◽  
Rrna Sequencing ◽  
Stool Samples ◽  
Microbiota Diversity

AbstractObjectiveThe aim of this study is to investigate the dysbiosis characteristics of gut microbiota in patients with cerebral infarction (CI) and its clinical implications.MethodsStool samples were collected from 79 CI patients and 98 healthy controls and subjected to 16S rRNA sequencing to identify stool microbes. Altered compositions and functions of gut microbiota in CI and its correlation with clinical features were investigated. Random forest and receiver operating characteristic analysis were used to develop a diagnostic model.ResultsMicrobiota diversity and structure between CI patients and healthy controls were overall similar. However, butyrate-producing bacteria (BPB) were significantly reduced in CI patients, while lactic acid bacteria (LAB) were increased. Genetically, BPB-related functional genes were reduced in CI patients, whereas LAB-related genes were enhanced. The interbacterial correlations among BPB in CI patients were less prominent than those in healthy controls. Clinically, BPB was negatively associated with the National Institutes of Health Stroke Scale (NIHSS), while LAB was positively correlated with NIHSS. Both BPB and LAB played leading roles in the diagnostic model based on 47 bacteria.ConclusionsThe abundance and functions of BPB in CI patients were significantly decreased, while LAB were increased. Both BPB and LAB displayed promising potential in the assessment and diagnosis of CI.

The evaluation of physiologic decannulation readiness according to upper airway resistance measurement

2008 ◽  
Vol 139 (4) ◽  
pp. 535-540 ◽  
Author(s):  
Chunli Gao ◽  
Liang Zhou ◽  
Chunsheng Wei ◽  
Matthew R. Hoffman ◽  
Cai Li ◽  
...  
Keyword(s):  
Airway Resistance ◽  
Operating Characteristic ◽  
Upper Airway ◽  
High Sensitivity ◽  
Deep Breathing ◽  
Clinical Criteria ◽  
Receiver Operating Characteristic Curves ◽  
Upper Airway Resistance ◽  
Laryngeal Disease ◽  
Subglottal Pressure

Objective To measure the upper-airway resistance in patients with tracheostomies and determine the value representing decannulation readiness. Subjects and Methods Fifty-six patients with tracheostomies resultant to laryngeal disease participated in this study. Forty patients met clinical criteria for decannulation; 16 did not. Subglottal pressure was measured with a tube connected to the tracheostomy tube, and airflow was monitored simultaneously using a facemask. Upper-airway resistance measurements were recorded during shallow and deep breathing. Results During both shallow and deep breathing, the inspiratory and expiratory resistances were significantly higher for the group unsuitable for decannulation ( P < .0001). The areas under the receiver operating characteristic curves were 0.938 or greater for the four curves, indicating a high sensitivity and specificity of resistance measures for diagnosis. Conclusions Objective measurement of upper-airway resistance during shallow and deep breathing may be a useful parameter in determining decannulation readiness of tracheostomized patients.

Diagnostic and prognostic significance of miR-451a in patients with atherosclerosis

Vascular ◽  
2021 ◽  
pp. 170853812110585
Author(s):  
Baizhi Wang ◽  
Xingliang Duan ◽  
Qing Xu ◽  
Yani Li
Keyword(s):  
Operating Characteristic ◽  
Cox Regression ◽  
Prognostic Biomarker ◽  
Prognostic Significance ◽  
High Sensitivity ◽  
Roc Curves ◽  
Cox Regression Analysis ◽  
Chi Squared ◽  
Cardiovascular Endpoint ◽  
Inflammatory Vascular Disease

Objectives Atherosclerosis (AS) is a chronic inflammatory vascular disease. This study aimed to detect the expression level of miR-451a and investigate the diagnostic and prognostic values of miR-451a for AS patients. Methods The relative expression of miR-451a was assessed by qRT-PCR. Comparison of groups was analyzed with the t-test and chi-squared test. Pearson analysis was used to validate the correlation of miR-451 with CRP and CIMT. The receiver operating characteristic (ROC) curves, K-M analysis, and Cox regression analysis were conducted to explore the roles of miR-451a in diagnosing AS patients and predicting outcomes of AS patients. Results The expression of miR-451a was significantly decreased in the serum of AS patients. The results of Pearson analysis showed the expression of miR-451a was negatively correlated with CRP and CIMT. The data of ROC proposed miR-451a could differentiate AS patients from healthy individuals with high sensitivity and specificity. K-M analysis and Cox regression showed miR-451a might be an independent biomarker of suffering cardiovascular endpoint diseases in AS patients. The expression of miR-451a was obviously inhibited in AS patients with cardiovascular endpoint events. Conclusion Deregulation of miR-451a might be associated with the development of AS. MiR-451a might be used as a promising diagnostic and prognostic biomarker for clinical treatment of AS patients.

scholarly journals Alzheimer’s Disease and Parkinson Dementia Distinguished by Cognitive Marker

2020 ◽  
Author(s):  
Irina Kozlova ◽  
Mario A Parra ◽  
Nataliya Titova ◽  
Maria Gantman ◽  
Sergio Della Sala
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Cognitive Impairment ◽  
Operating Characteristic ◽  
Characteristic Curve ◽  
High Sensitivity ◽  
Task Demands ◽  
Memory Binding ◽  
Parkinson Dementia ◽  
Temporary Memory

Abstract Background Temporary memory binding (TMB) has been shown to be specifically affected by Alzheimer’s disease (AD) when it is assessed via free recall and titrating the task demands to equate baseline performance across patients. Methods Patients with Parkinson’s disease (PD) were subdivided into patients with and without cognitive impairment and compared with AD and amnestic mild cognitive impairment (aMCI) patients on their performance on the TMB. Results The results show that only patients with AD dementia present with impaired TMB performance. Receiver operating characteristic curve analyses showed that TMB holds high sensitivity and specificity for aMCI and AD relative to PD groups and healthy controls. Conclusion The TMB is sensitive to the neurodegenerative mechanisms leading to AD dementia but not to those underpinning PD dementia. As such, TMB task can aid the differential diagnosis of these common forms of dementia.

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