scholarly journals Development and Validation of a Novel Leishmania donovani Screening Cascade for High-Throughput Screening Using a Novel Axenic Assay with High Predictivity of Leishmanicidal Intracellular Activity

2015 ◽  
Vol 9 (9) ◽  
pp. e0004094 ◽  
Author(s):  
Andrea Nühs ◽  
Manu De Rycker ◽  
Sujatha Manthri ◽  
Eamon Comer ◽  
Christina A. Scherer ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Leishmania Donovani ◽  
Intracellular Activity ◽  
Development And Validation ◽  
Screening Cascade

scholarly journals Use of Leishmania donovani Field Isolates Expressing the Luciferase Reporter Gene in In Vitro Drug Screening

2005 ◽  
Vol 49 (9) ◽  
pp. 3776-3783 ◽  
Author(s):  
Ashutosh ◽  
Suman Gupta ◽  
Ramesh ◽  
Shyam Sundar ◽  
Neena Goyal
Keyword(s):  
Reporter Gene ◽  
High Throughput ◽  
High Throughput Screening ◽  
Leishmania Donovani ◽  
Luciferase Activity ◽  
Firefly Luciferase ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Field Isolates

ABSTRACT Currently available primary screens for the selection of candidate antileishmanial compounds are not ideal. These techniques are time-consuming, laborious, and difficult to scale and require macrophages, which limit their use for high-throughput screening. We have developed Leishmania donovani field isolates that constitutively express the firefly luciferase reporter gene (luc) as a part of an episomal vector. An excellent correlation between parasite number and luciferase activity was observed. luc expression was stable, even in the absence of drug selection, for 4 weeks. The transfectants were infective to macrophages, and intracellular amastigotes exhibited luciferase activity. The suitability of these recombinant field isolates for in vitro screening of antileishmanial drugs was established. The luciferase-expressing sodium stibogluconate-resistant cell lines offer a model for the screening of compounds for resistance. The system is in routine use at the Central Drug Research Institute, Lucknow, India, for high-throughput screening of newly synthesized compounds.

Development and validation of high-throughput screening assays for poly(ADP-ribose) polymerase-2 inhibitors

2014 ◽  
Vol 449 ◽  
pp. 188-194 ◽  
Author(s):  
Zhixiang Zhu ◽  
Jing Jin ◽  
Nina Xue ◽  
Xiuyun Song ◽  
Xiaoguang Chen
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Screening Assays ◽  
Development And Validation

scholarly journals Development of a High-Throughput Cytometric Screen to Identify Anti-Wolbachia Compounds: The Power of Public–Private Partnership

2019 ◽  
Vol 24 (5) ◽  
pp. 537-547
Author(s):  
Rachel H. Clare ◽  
Roger Clark ◽  
Catherine Bardelle ◽  
Paul Harper ◽  
Matthew Collier ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Neglected Tropical Diseases ◽  
Bacterial Symbiont ◽  
Tropical Diseases ◽  
Validation Process ◽  
Host Insect ◽  
Filarial Nematodes ◽  
Development And Validation ◽  
High Throughput Assay

The Anti- Wolbachia (A·WOL) consortium at the Liverpool School of Tropical Medicine (LSTM) has partnered with the Global High-Throughput Screening (HTS) Centre at AstraZeneca to create the first anthelmintic HTS for neglected tropical diseases (NTDs). The A·WOL consortium aims to identify novel macrofilaricidal drugs targeting the essential bacterial symbiont ( Wolbachia) of the filarial nematodes causing onchocerciasis and lymphatic filariasis. Working in collaboration, we have validated a robust high-throughput assay capable of identifying compounds that selectively kill Wolbachia over the host insect cell. We describe the development and validation process of this complex, phenotypic high-throughput assay and provide an overview of the primary outputs from screening the AstraZeneca library of 1.3 million compounds.

scholarly journals Development and validation of a hedgehog heparin-binding assay for high-throughput screening

MethodsX ◽  
2020 ◽  
pp. 101207
Author(s):  
David Lamson ◽  
Mark Hughes ◽  
Audrey Adcock ◽  
Ginger Smith ◽  
Kevin P. Williams
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Binding Assay ◽  
Heparin Binding ◽  
Development And Validation

scholarly journals Development of a Novel Ectonucleotidase Assay Suitable for High-Throughput Screening

2012 ◽  
Vol 17 (7) ◽  
pp. 993-998 ◽  
Author(s):  
Kris F. Sachsenmeier ◽  
Carl Hay ◽  
Erin Brand ◽  
Lori Clarke ◽  
Kim Rosenthal ◽  
...  
Keyword(s):  
Malachite Green ◽  
High Throughput ◽  
High Throughput Screening ◽  
High Performance ◽  
Adenosine Monophosphate ◽  
Screening Assay ◽  
Highly Sensitive ◽  
Development And Validation ◽  
Multiple Samples

5′-Ectonucleotidase (NT5E) catalyzes the conversion of adenosine monophosphate to adenosine and free phosphate. The role of this ectonucleotidase and its production of adenosine are linked with immune function, angiogenesis, and cancer. NT5E activity is typically assayed either by chromatographic quantification of substrates and products using high-performance liquid chromatography (HPLC) or by quantification of free phosphate using malachite green. These methods are not suitable for robust screening assays of NT5E activity. HPLC is not readily suitable for the rapid and efficient assay of multiple samples and malachite green is highly sensitive to the phosphate-containing buffers common in various media and sample buffers. Here the development and validation of a novel high-throughput ectonucleotidase screening assay are described, which makes use of a luciferase-based assay reagent, the Promega CellTiter-Glo kit, to measure the catabolism of AMP by NT5E. This multiwell plate-based assay facilitates the screening of potential ectonucleotidase antagonists and is unaffected by the presence of contaminating phosphate molecules present in screening samples.

Sign in / Sign up

Export Citation Format

Share Document