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Assays Research

12:03 EDT 19th August 2018 | BioPortfolio

Cell Penetration Profiling Using the Chloroalkane Penetration Assay.

Biotherapeutics are a promising class of molecules in drug discovery, but they are often limited to extracellular targets due to their poor cell penetration. High-throughput cell penetration assays are required for the optimization of biotherapeutics for enhanced cell penetration. We developed a HaloTag-based assay called the ChloroAlkane Penetration Assay (CAPA), which is quantitative, high-throu...

Development of a TaqMan-based real-time PCR for detecting duck adenovirus 3.

Recently, a novel duck adenovirus (designated as duck adenovirus 3, DAdV-3) was discovered in Muscovy ducks, China. Here, we developed a TaqMan-based real-time PCR assay (qPCR) for the detection of DAdV-3 infection. After the optimization of the qPCR conditions, the lower limit of detection for DAdV-3 infection was 40.9 copies/μl. No cross-reactivity was observed with other duck-derived pathogens...

Perspective: SBI at Its Fifth Annual Meeting.

HCS Campaign to Identify Compounds That Inhibit or Disrupt Androgen Receptor-Transcriptional Intermediary Factor 2 Protein-Protein Interactions for the Treatment of Prostate Cancer.

Twenty percent of prostate cancer (PCa) patients develop a noncurable drug-resistant form of the disease termed castration-resistant prostate cancer (CRPC). Overexpression of Androgen Receptor (AR) coactivators such as transcriptional intermediary factor 2 (TIF2) is associated with poor CRPC patient outcomes. We describe the implementation of the AR-TIF2 protein-protein interaction biosensor (PPIB...

A diagnostic evaluation of a molecular assay used for testing and treating anorectal chlamydia and gonorrhoea infections at the point-of-care in Papua New Guinea.

Papua New Guinea (PNG) has among the highest prevalences of sexually transmissible infections (STIs) globally with no services able to accurately test for anorectal Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) infections. Here we prospectively evaluated the diagnostic performance of a molecular CT/NG assay used at the point-of-care (POC) with the aim of enhancing anorectal STI screeni...

A real time high-resolution melting PCR assay for detection and differentiation among sheep pox virus, goat pox virus, field isolates and vaccine strains of lumpy skin disease virus.

In this paper, we report on the development of a real time high-resolution melting (HRM) PCR assay for detection and differentiation among sheep pox virus (SPPV), goat pox virus (GTPV), field isolates and vaccine strains of lumpy skin disease virus (LSDV) based on high-resolution melting curve analysis of their target PCR amplicons. A 111 bp region of LSDV010 ORF, which harbors unique genetic diff...

Small-Molecule Screening Assay for Mono-ADP-Ribosyltransferases.

Mono-ADP-ribosyltransferases of the PARP/ARTD enzyme family are enzymes catalyzing the transfer of a single ADP-ribose unit to target proteins. The enzymes have various roles in vital cellular processes such as DNA repair and transcription, and many of the enzymes are linked to cancer-relevant functions. Thus inhibition of the enzymes is a potential way to discover and develop new drugs against ca...

A Simple, Sensitive, and Generalizable Plate Assay for Screening PARP Inhibitors.

Poly-ADP-ribose polymerases (also known as ADP-ribosyltransferases or ARTDs) are a family of 17 enzymes in humans that catalyze the reversible posttranslational modification known as ADP-ribosylation. PARPs are implicated in diverse cellular processes, from DNA repair to the unfolded protein response. Small-molecule inhibitors of PARPs have improved our understanding of PARP-mediated biology and, ...

Pyrazinyl ureas revisited: 1-(3-(Benzyloxy)pyrazin-2-yl)-3-(3,4-dichlorophenyl)urea, a new blocker of Aβ-induced mPTP opening for Alzheimer's disease.

Herein, we report synthesis and evaluation of new twenty-eight pyrazinyl ureas against β amyloid (Aβ)-induced opening of mitochondrial permeability transition pore (mPTP) using JC-1 assay which measures the change of mitochondrial membrane potential (ΔΨm). The neuroprotective effect of seventeen compounds against Aβ-induced mPTP opening was superior to that of the standard Cyclosporin A (CsA)...

Development of a real time loop-mediated isothermal amplification method for detection of Senecavirus A.

Senecavirus A (SVA), formerly known as Seneca Valley Virus (SVV), is one of causative agents of vesicular diseases in swine. Recently, the outbreaks associated with vesicular disease caused by SVA infection in pig herds have been reported in Brazil, USA, China, Thailand and Canada. Several molecular detection methods have been established to determine the infection of SVA, including real time reve...

Quantitative detection of hepatitis C virus RNA in urine of patients with chronic hepatitis C using a novel real-time PCR assay.

Hepatitis C virus (HCV) RNA can be detected in body fluids such as urine. However, because of deficiencies in established isolation and detection methods, the actual prevalence and form of HCV RNA in the urine of patients with hepatitis C remain unclear. To more sensitively and accurately measure urine HCV RNA levels, a novel real-time PCR assay with a modified isolation method and short amplicon ...

A triplex probe-based TaqMan qPCR assay for Calreticulin type I and II mutation detection.

Calreticulin (CALR) exon 9 frameshift mutations have recently been identified in 30-40% of patients with essential thrombocythemia (ET) and primary myelofibrosis (PMF) without JAK2 or MPL mutations. We aimed to develop a qPCR assay to screen type I and II mutations of CALR.

Multicenter evaluation of the Alere™ i influenza A&B assay using respiratory specimens collected in viral transport media.

Rapid and accurate detection of influenza virus is critical for proper patient management. The Alere™ i Influenza A&B assay is an isothermal nucleic acid amplification test capable of detecting influenza A and B viruses directly from respiratory specimens. In this multicenter clinical trial conducted in the US, we evaluated the clinical performance of the Alere™ i Influenza A&B assay against t...

Sensitive detection of live Escherichia coli by bacteriophage amplification-coupled immunoassay on the Luminex® MAGPIX instrument.

Phages are natural predators of bacteria and have been exploited in bacterial detection because of their exquisite specificity to their cognate bacterial hosts. In this study, we present a "proof of concept" bacteriophage amplification-coupled assay as a surrogate for detecting a bacterium present in a sample. The assay entails detection of progeny phage resulting from infection and subsequent gro...

Development of a high throughput yeast-based screening assay for human carbonic anhydrase isozyme II inhibitors.

Carbonic anhydrase (CA; EC 4.2.1.1) catalyzes the reversible hydration of carbon dioxide (CO) to bicarbonate and proton. There are 16 known isozymes of α-CA in humans, which differ widely in their kinetics, subcellular localization and tissue-specific distribution. Several disorders are associated with abnormal levels of CA, and so the inhibition of CA has pharmacological application in the treat...

A TaqMan-based real-time PCR assay for the detection of Ungulate bocaparvovirus 2.

Ungulate bocaparvoviruses (UBoV) 2-5 are recently discovered porcine bocaparvoviruses belonging to the family Parvoviridae, and are considered to be a potentially major cause of swine diseases. In order to detect local UBoV2 epidemics in China, we developed a TaqMan-based real-time PCR (qPCR) assay targeting the UBoV2 VP1 gene and compared the results of qPCR with conventional PCR (cPCR). The qPCR...

Multicomponent spectrofluorimetric method for the assay of formylmethylflavin and its hydrolytic products: Kinetic applications.

A multicomponent spectrofluorimetric method has been developed for the simultaneous assay of formylmethylflavin (FMF), an intermediate product in the photolysis of riboflavin (vitamin B), and its side-chain hydrolytic products, lumichrome (LC) in acidic solution and LC and lumiflavin (LF) in the alkaline solution as well as its ring cleavage products, 1,2-dihydro-1-methyl-2-keto-3-quinoxaline carb...

Quantitative screening for endocrine-disrupting bisphenol A in consumer and household products using NanoAptamer assay.

The NanoAptamer assay is a bisphenol A (BPA) quantification method that uses magnetic beads, quantum dot nanoparticles, and a BPA-specific aptamer. In this study, screening of various consumer and household products for BPA was demonstrated utilizing the NanoAptamer assay. First, the experimental conditions suitable for BPA detection using the NanoAptamer assay were examined in terms of incubation...

Comparison of factors affecting platelet reactivity in various platelet function tests.

Previous studies have reported that various factors affect ADP-induced platelet reactivity during clopidogrel therapy. The aim of this study was to determine whether clinical and laboratory variables for platelet reactivity during dual antiplatelet therapy (DAPT) are dependent on the assay used. We enrolled 904 patients receiving DAPT following coronary intervention. Platelet reactivity was measur...

Development of a robust and standardized immunoserological assay for detection of anti-measles IgG antibodies in human sera.

Because of measles outbreaks there is a need for continuous monitoring of immunological protection against infection at population level. For such monitoring to be feasible, a cost-effective, reliable and high-throughput assay is necessary. Herein we describe an ELISA protocol for assessment of anti-measles antibody levels in human serum samples that fulfills the above criteria and is easily adapt...

Binding of host factors to stabilized HIV-1 capsid tubes.

The capsid-binding assay is an in vitro experiment used to determine whether cellular proteins interact with the HIV-1 core. In vitro assembled HIV-1 capsids recapitulate the surface of the HIV-1 core. The assay involves the incubation of in vitro assembled HIV-1 capsid-nucleocapsid (CA-NC) complexes with the protein in question. Subsequently, the mixture is spun through a sucrose cushion using an...

Assessment of the skin sensitisation hazard of functional polysiloxanes and silanes in the SENS-IS assay.

In the context of a larger testing programme that aimed at assessing the skin sensitisation potential of functional polysiloxanes and silanes, this investigation complements the available in vitro and in vivo data with data in the SENS-IS assay, a human in vitro 3D skin-based model. The SENS-IS assay allowed testing of all functional polysiloxanes and silanes without any solubility issues or limit...

Signal amplification by strand displacement in a carbon dot based fluorometric assay for ATP.

A fluorometric ATP assay is described that makes use of carbon dots and graphene oxide along with toehold-mediated strand displacement reaction. In the absence of target, the fluorescence of carbon dots (with excitation/emission maxima at 360/447 nm) is strong and in the "on" state, because the signal probe hybridizes with the aptamer strand and cannot combine with graphene oxide. In the presence...

Evaluation and validation of HPV real-time PCR assay for the detection of HPV DNA in oral cytobrush and FFPE samples.

Specific HPV genotypes have been recognized as risk factors inducing head and neck cancers (HNC). The aim of this study was to validate a real-time PCR assay to detect accurately High Risk HPV DNA in Formalin Fixed Paraffin Embedded (FFPE) and oral cytobrush samples and compare the results with conventional PCR. Repeatability, reproducibility and limit of detection of Cobas assay were estimated fo...

A tailored reverse transcription loop-mediated isothermal amplification for sensitive and specific detection of serotype A foot-and-mouth disease virus circulating in pool 1 region countries.

Rapid and accurate diagnosis of foot-and-mouth disease viruses (FMDV) is essential for the prompt control of FMD outbreaks. Reverse transcription polymerase chain reaction (RT-PCR) and real-time quantitative RT-PCR (qRT-PCR) are used for routine FMDV diagnosis as World Organisation for Animal Health-recommended diagnostic assays. However, these PCR-based assays require sophisticated equipment, spe...

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