scholarly journals 1088. Ultrasensitive Detection of C. difficile Toxins in Stool Using Single Molecule Counting Technology: A Multicenter Study for Evaluation of Clinical Performance

2018 ◽  
Vol 5 (suppl_1) ◽  
pp. S325-S326 ◽  
Author(s):  
Stephen Young ◽  
Ray Mills ◽  
Christen Griego-Fullbright ◽  
Aaron Wagner ◽  
Emily Herding ◽  
...  
Keyword(s):  
Single Molecule ◽  
Multicenter Study ◽  
Clinical Performance ◽  
Turnaround Time ◽  
Neutralization Assay ◽  
Single Step ◽  
Reporting Structure ◽  
One Step ◽  
Automated Immunoassay ◽  
Higher Sensitivity

Abstract Background Commercially available tests for Clostridium difficile infection (CDI) make test selection by the laboratory difficult due to the following unsatisfactory characteristics: long turnaround time, poor sensitivity, and/or poor specificity. The Singulex Clarity® C. diff toxins A/B assay (in development) is a rapid and automated immunoassay for the detection of C. difficile toxins A and B in stool, with analytical limits of detection for toxins A and B at 2.0 and 0.7 pg/mL, respectively. In this multicenter study, the clinical performance of the Singulex Clarity C. diff toxins A/B assay was compared with standalone PCR, a multistep algorithm with enzyme immunoassay (EIA) and PCR, and cell cytotoxicity neutralization assay (CCNA). Methods Fresh samples from 267 subjects with suspected CDI were tested at two sites (Minneapolis Medical Research Foundation and TriCore Reference Laboratories) with the Singulex Clarity assay, PCR (Xpert®C. difficile), and EIA (C. Diff Quik Chek Complete®) for GDH and toxin testing. The performance of the assays and multistep algorithms were evaluated against CCNA (Microbiology Specialists, Inc.). Results The overall CDI prevalence was 15.7%. The Singulex Clarity C. diff toxins A/B assay had 90.5% sensitivity and 96.0% specificity, with a 98.2% negative predictive value when compared with CCNA, and the Clarity assay’s AuROC was 0.9534. PCR had 90.5% sensitivity and 91.1% specificity. Compared with CCNA, the toxin EIA had 47.6% sensitivity and 100% specificity. Testing with a multistep algorithm using EIA with discordant results reflexed to PCR resulted in 85.7% sensitivity and 94.7% specificity. Conclusion The ultrasensitive Singulex Clarity C. diff toxins A/B assay is equivalent to the sensitivity of PCR while providing higher specificity. Compared with a multistep algorithm, the Clarity assay provides higher sensitivity and specificity while providing faster time-to-result in a simpler-to-understand, one-step reporting structure, allowing for a standalone, single-step solution for detection of C. difficile toxins in patients with suspected CDI. Disclosures E. Friedland, Singulex, Inc.: Employee, Salary. A. Bartolome, Singulex, Inc.: Employee, Salary. A. Almazan, Singulex, Inc.: Employee, Salary. S. Tam, Singulex, Inc.: Employee, Salary. S. Biscocho, Singulex, Inc.: Employee, Salary. S. Abusali, Singulex, Inc.: Employee, Salary. J. Sandlund, Singulex, Inc.: Employee, Salary. J. Estis, Singulex, Inc.: Employee, Salary. J. Bishop, Singulex, Inc.: Employee, Salary.

scholarly journals Increased Clinical Specificity with Ultrasensitive Detection of Clostridioides difficile Toxins: Reduction of Overdiagnosis Compared to Nucleic Acid Amplification Tests

2019 ◽  
Vol 57 (11) ◽  
Author(s):  
Johanna Sandlund ◽  
Joel Estis ◽  
Phoebe Katzenbach ◽  
Niamh Nolan ◽  
Kirstie Hinson ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Patient Outcomes ◽  
Single Molecule ◽  
Clinical Performance ◽  
Neutralization Assay ◽  
Nucleic Acid Amplification ◽  
Content Type ◽  
Clostridioides Difficile ◽  
Health Care Associated Infections ◽  
Stool Specimens

ABSTRACT Clostridioides difficile infection (CDI) is one of the most common health care-associated infections, resulting in significant morbidity, mortality, and economic burden. Diagnosis of CDI relies on the assessment of clinical presentation and laboratory tests. We evaluated the clinical performance of ultrasensitive single-molecule counting technology for detection of C. difficile toxins A and B. Stool specimens from 298 patients with suspected CDI were tested with the nucleic acid amplification test (NAAT; BD MAX Cdiff assay or Xpert C. difficile assay) and Singulex Clarity C. diff toxins A/B assay. Specimens with discordant results were tested with the cell cytotoxicity neutralization assay (CCNA), and the results were correlated with disease severity and outcome. There were 64 NAAT-positive and 234 NAAT-negative samples. Of the 32 NAAT+/Clarity− and 4 NAAT−/Clarity+ samples, there were 26 CCNA− and 4 CCNA− samples, respectively. CDI relapse was more common in NAAT+/toxin+ patients than in NAAT+/toxin− and NAAT−/toxin− patients. The clinical specificity of Clarity and NAAT was 97.4% and 89.0%, respectively, and overdiagnosis was more than three times more common in NAAT+/toxin− than in NAAT+/toxin+ patients. The Clarity assay was superior to NAATs for the diagnosis of CDI, by reducing overdiagnosis and thereby increasing clinical specificity, and the presence of toxins was associated with negative patient outcomes.

scholarly journals Ultrasensitive Detection of Clostridioides difficile Toxins in Stool by Use of Single-Molecule Counting Technology: Comparison with Detection of Free Toxin by Cell Culture Cytotoxicity Neutralization Assay

2019 ◽  
Vol 57 (11) ◽  
Author(s):  
Glen Hansen ◽  
Stephen Young ◽  
Alan H. B. Wu ◽  
Emily Herding ◽  
Vickie Nordberg ◽  
...  
Keyword(s):  
Cell Culture ◽  
Single Molecule ◽  
Neutralization Assay ◽  
Single Step ◽  
Multicenter Studies ◽  
Content Type ◽  
Clostridioides Difficile ◽  
Ultrasensitive Assay ◽  
A Cell ◽  
Positive Agreement

ABSTRACT Laboratory tests for Clostridioides difficile infection (CDI) rely on the detection of free toxin or molecular detection of toxin genes. The Singulex Clarity C. diff toxins A/B assay is a rapid, automated, and ultrasensitive assay that detects C. difficile toxins A and B in stool. We compared CDI assays across two prospective multicenter studies to set a cutoff for the Clarity assay and to independently validate the performance compared with that of a cell culture cytotoxicity neutralization assay (CCCNA). The cutoff was set by two sites testing fresh samples from 897 subjects with suspected CDI and then validated at four sites testing fresh samples from 1,005 subjects with suspected CDI. CCCNA testing was performed at a centralized laboratory. Samples with discrepant results between the Clarity assay and CCCNA were retested with CCCNA when the Clarity result agreed with that of at least one comparator method; toxin enzyme immunoassays (EIA), glutamate dehydrogenase (GDH) detection, and PCR were performed on all samples. The cutoff for the Clarity assay was set at 12.0 pg/ml. Compared to results with CCCNA, the Clarity assay initially had 85.2% positive agreement and 92.4% negative agreement. However, when samples with discrepant results between the Clarity assay and CCCNA in the validation study were retested by CCCNA, 13/17 (76.5%) Clarity-negative but CCCNA-positive samples (Clarity+/CCCNA−) became CCCNA−, and 5/26 (19.2%) Clarity+/CCCNA− samples became CCCNA+, resulting in a 96.3% positive agreement and 93.0% negative agreement between Clarity and CCCNA results. The toxin EIA had 59.8% positive agreement with CCCNA. The Clarity assay was the most sensitive free-toxin immunoassay, capable of providing CDI diagnosis in a single-step solution. A different CCCNA result was reported for 42% of retested samples, increasing the positive agreement between Clarity and CCCNA from 85.2% to 96.3% and indicating the challenges of comparing free-toxin results to CCCNA results as a reference standard.

scholarly journals 2356. Increased Clinical Specificity with Ultrasensitive Detection of Clostridioides difficile Toxins: Reduction of Overdiagnosis Compared with Nucleic Acid Amplification Tests

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S811-S812 ◽  
Author(s):  
Johanna Sandlund ◽  
Joel Estis ◽  
Phoebe Katzenbach ◽  
Niamh Nolan ◽  
Kirstie Hinson ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Disease Severity ◽  
Single Molecule ◽  
Clinical Performance ◽  
Neutralization Assay ◽  
Nucleic Acid Amplification ◽  
Clostridioides Difficile ◽  
Percent Agreement ◽  
Clostridioides Difficile Infection ◽  
Healthcare Associated

Abstract Background Clostridioides difficile infection (CDI) is one of the most common healthcare-associated infections, resulting in significant morbidity, mortality, and economic burden. Diagnosis of CDI relies on the assessment of clinical presentation and laboratory tests. We have evaluated the clinical performance of ultrasensitive Single Molecule Counting technology for detection of C. difficile toxins A and B. Methods Stool specimens from 298 patients with suspected CDI were tested with nucleic acid amplification test (NAAT; BD MAX™ Cdiff assay or Xpert® C. difficile assay) and Singulex Clarity® C. difficile toxins A/B assay. Specimens with discordant results were tested with cell cytotoxicity neutralization assay (CCNA), and results were correlated with disease severity and outcome. Results There were 64 NAAT-positive and 234 NAAT-negative samples. Of the 32 NAAT+/Clarity− and 4 NAAT-/Clarity+ samples, there were 26 CCNA− and 4 CCNA- samples, respectively. CDI relapse or overall death was more common in NAAT+/toxin+ patients than in NAAT+/toxin− and NAAT−/toxin− patients, and NAAT+/toxin+ patients were 3.7 times more likely to experience relapse or death (Figure 1). The clinical specificity of Clarity and NAAT was 97.4% and 89.0%, respectively, and overdiagnosis was over three times more common in NAAT+/toxin− than in NAAT+/toxin+ patients (Figure 2). Negative percent agreement between NAAT and Clarity was 98.3%, and positive percent agreement increased from 50.0% to effective 84.2% and 94.1% after CCNA testing and clinical assessment. Conclusion The Clarity assay was superior to NAATs in diagnosis of CDI, by reducing overdiagnosis and thereby increasing clinical specificity, and presence of toxins was associated with disease severity and outcome. Disclosures All authors: No reported disclosures.

Synthesis of Polycycles and Oxacycles by Tandem Metathesis of endo–norbornene Derivatives

Synthesis ◽  
2021 ◽  
Author(s):  
Sambasivarao Kotha ◽  
Sunil Pulletikurti ◽  
Ambareen Fatma ◽  
gopal dhangar ◽  
gonna somu Naidu
Keyword(s):  
Natural Products ◽  
Metal Complex ◽  
Carbonyl Group ◽  
Single Step ◽  
Time Dependent ◽  
Computational Studies ◽  
Nmr Studies ◽  
Tricyclic Compounds ◽  
One Step ◽  
Norbornene Derivatives

Here, we have demonstrated that the presence of a carbonyl group at C7 position is preventing the olefin metathesis of endo-norbornene derivatives due to the complexation of the metal alkylidene. Time-dependent NMR studies showed the presence of new proton signals in the metal alkylidene region, which indicate the formation of metal complex with the carbonyl group of the substrate. These observations were further proved by ESI-MS analysis. Whereas, computational studies provided that the catalyst was interacting with the C7 carbonyl group and aligned perpendicular to that of norbornene olefin. Later, these endo-keto norbornene derivatives were reduced to hydroxyl derivatives diastereoselectively. Ring-rearrangement metathesis (RRM) of these hydroxyl derivatives, produced the [6/5/6], and [5/6/5] carbo-tricyclic cores of the natural products in one step. Whereas the RRM of O-allyl derivatives, delivered the oxa-tricyclic compounds in a single step with excellent yields.

scholarly journals One-Year Clinical Evaluation of the Bonding Effectiveness of a One-Step, Self-Etch Adhesive in Noncarious Cervical Lesion Therapy

2015 ◽  
Vol 2015 ◽  
pp. 1-5 ◽  
Author(s):  
Babacar Faye ◽  
Mouhamed Sarr ◽  
Khaly Bane ◽  
Adjaratou Wakha Aidara ◽  
Seydina Ousmane Niang ◽  
...  
Keyword(s):  
Clinical Performance ◽  
Retention Rate ◽  
Recall Rate ◽  
Acid Etching ◽  
Significant Difference ◽  
Usphs Criteria ◽  
One Step ◽  
One Year ◽  
The One ◽  
Self Etch

This study evaluated the one-year clinical performance of a one-step, self-etch adhesive (Optibond All-in-One, Kerr, CA, USA) combined with a composite (Herculite XRV Ultra, Kerr Hawe, CA, USA) to restore NCCLs with or without prior acid etching. Restorations performed by the same practitioner were evaluated at baseline and after 3, 6, and 12 months using modified USPHS criteria. At 6 months, the recall rate was 100%. The retention rate was 84.2% for restorations with prior acid etching, but statistically significant differences were observed between baseline and 6 months. Without acid etching, the retention rate was 77%, and no statistically significant difference was noted between 3 and 6 months. Marginal integrity (93.7% with and 87.7% without acid etching) and discoloration (95.3% with and 92.9% without acid etching) were scored as Alpha or Bravo, with better results after acid etching. After one year, the recall rate was 58.06%. Loss of pulp vitality, postoperative sensitivity, or secondary caries were not observed. After one year retention rate was of 90.6% and 76.9% with and without acid conditioning. Optibond All-in-One performs at a satisfactory clinical performance level for restoration of NCCLs after 12 months especially after acid etching.

scholarly journals Single Step 2-Port Device De-Embedding Algorithm for Fixture-DUT-Fixture Network Assembly

Electronics ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 1275
Author(s):  
Simone Scafati ◽  
Enza Pellegrino ◽  
Francesco de Paulis ◽  
Carlo Olivieri ◽  
James Drewniak ◽  
...  
Keyword(s):  
Standard Technique ◽  
Single Step ◽  
Scattering Parameters ◽  
Linear Set ◽  
S Parameter ◽  
Parameter Conversion ◽  
Before And After ◽  
One Step ◽  
Typical Measurement

The de-embedding of measurement fixtures is relevant for an accurate experimental characterization of radio frequency and digital electronic devices. The standard technique consists in removing the effects of the measurement fixtures by the calculation of the transfer scattering parameters (T-parameters) from the available measured (or simulated) global scattering parameters (S-parameters). The standard de-embedding is achieved by a multiple steps process, involving the S-to-T and subsequent T-to-S parameter conversion. In a typical measurement setup, two fixtures are usually placed before and after the device under test (DUT) allowing the connection of the device to the calibrated vector network analyzer coaxial ports. An alternative method is proposed in this paper: it is based on the newly developed multi-network cascading algorithm. The matrices involved in the fixture-DUT-fixture cascading gives rise to a non-linear set of equations that is in one step analytically solved in closed form, obtaining a unique solution. The method is shown to be effective and at least as accurate as the standard multi-step de-embedding one.

scholarly journals Computational and experimental investigations of one-step conversion of poly(carbonate)s into value-added poly(aryl ether sulfone)s

2016 ◽  
Vol 113 (28) ◽  
pp. 7722-7726 ◽  
Author(s):  
Gavin O. Jones ◽  
Alexander Yuen ◽  
Rudy J. Wojtecki ◽  
James L. Hedrick ◽  
Jeannette M. García
Keyword(s):  
High Performance ◽  
Density Functional ◽  
Value Added ◽  
Single Step ◽  
Nucleophilic Attack ◽  
Experimental Investigations ◽  
Aryl Ether ◽  
One Step ◽  
Poly Carbonate

It is estimated that ∼2.7 million tons poly(carbonate)s (PCs) are produced annually worldwide. In 2008, retailers pulled products from store shelves after reports of bisphenol A (BPA) leaching from baby bottles, reusable drink bottles, and other retail products. Since PCs are not typically recycled, a need for the repurposing of the PC waste has arisen. We report the one-step synthesis of poly(aryl ether sulfone)s (PSUs) from the depolymerization of PCs and in situ polycondensation with bis(aryl fluorides) in the presence of carbonate salts. PSUs are high-performance engineering thermoplastics that are commonly used for reverse osmosis and water purification membranes, medical equipment, as well as high temperature applications. PSUs generated through this cascade approach were isolated in high purity and yield with the expected thermal properties and represent a procedure for direct conversion of one class of polymer to another in a single step. Computational investigations performed with density functional theory predict that the carbonate salt plays two important catalytic roles in this reaction: it decomposes the PCs by nucleophilic attack, and in the subsequent polyether formation process, it promotes the reaction of phenolate dimers formed in situ with the aryl fluorides present. We envision repurposing poly(BPA carbonate) for the production of value-added polymers.

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 A simple and versatile design concept for fluorophore derivatives with intramolecular photostabilization

2016 ◽  
Vol 7 (1) ◽  
Author(s):  
Jasper H. M. van der Velde ◽  
Jens Oelerich ◽  
Jingyi Huang ◽  
Jochem H. Smit ◽  
Atieh Aminian Jazi ◽  
...  
Keyword(s):  
Single Molecule ◽  
Unnatural Amino Acids ◽  
Photophysical Properties ◽  
Super Resolution ◽  
Single Step ◽  
General Strategy ◽  
Self Healing ◽  
Chemical Transformations ◽  
Scaffolding Strategy ◽  
Organic Fluorophore

Abstract Intramolecular photostabilization via triple-state quenching was recently revived as a tool to impart synthetic organic fluorophores with ‘self-healing’ properties. To date, utilization of such fluorophore derivatives is rare due to their elaborate multi-step synthesis. Here we present a general strategy to covalently link a synthetic organic fluorophore simultaneously to a photostabilizer and biomolecular target via unnatural amino acids. The modular approach uses commercially available starting materials and simple chemical transformations. The resulting photostabilizer–dye conjugates are based on rhodamines, carbopyronines and cyanines with excellent photophysical properties, that is, high photostability and minimal signal fluctuations. Their versatile use is demonstrated by single-step labelling of DNA, antibodies and proteins, as well as applications in single-molecule and super-resolution fluorescence microscopy. We are convinced that the presented scaffolding strategy and the improved characteristics of the conjugates in applications will trigger the broader use of intramolecular photostabilization and help to emerge this approach as a new gold standard.

scholarly journals TEMPO-Functionalized Nanoporous Au Nanocomposite for the Electrochemical Detection of H2O2

2018 ◽  
Vol 2018 ◽  
pp. 1-11
Author(s):  
Dongxiao Wen ◽  
Qianrui Liu ◽  
Ying Cui ◽  
Huaixia Yang ◽  
Jinming Kong
Keyword(s):  
Electrochemical Detection ◽  
Nanoporous Gold ◽  
Carboxyl Groups ◽  
Time Interval ◽  
Optimal Conditions ◽  
Sem Images ◽  
One Step ◽  
Ph Range ◽  
Synthesis Approach ◽  
Higher Sensitivity

A novel nanocomposite of nanoporous gold nanoparticles (np-AuNPs) functionalized with 2,2,6,6-tetramethyl-1-piperidinyloxy radical (TEMPO) was prepared; assembled carboxyl groups on gold nanoporous nanoparticles surface were combined with TEMPO by the “bridge” of carboxylate-zirconium-carboxylate chemistry. SEM images and UV-Vis spectroscopies of np-AuNPs indicated that a safe, sustainable, and simplified one-step dealloying synthesis approach is successful. The TEMPO-np-AuNPs exhibited a good performance for the electrochemical detection of H2O2 due to its higher number of electrochemical activity sites and surface area of 7.49 m2g-1 for load bigger amount of TEMPO radicals. The TEMPO-functionalized np-AuNPs have a broad pH range and shorter response time for H2O2 catalysis verified by the response of amperometric signal under different pH and time interval. A wide linear range with a detection limit of 7.8 × 10-7 M and a higher sensitivity of 110.403 μA mM-1cm-2 were obtained for detecting H2O2 at optimal conditions.

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