Development of a high-throughput assay for measuring lipase activity using natural triacylglycerols coated on microtiter plates

The Analyst ◽  
2013 ◽  
Vol 138 (18) ◽  
pp. 5230 ◽  
Author(s):  
Carole Serveau-Avesque ◽  
Robert Verger ◽  
Jorge A. Rodriguez ◽  
Abdelkarim Abousalham
Keyword(s):  
High Throughput ◽  
Lipase Activity ◽  
Microtiter Plates ◽  
High Throughput Assay

Development of a High-Throughput Assay for Measuring Phospholipase A Activity Using Synthetic 1,2-α-Eleostearoyl-sn-glycero-3-phosphocholine Coated on Microtiter Plates

2014 ◽  
Vol 86 (21) ◽  
pp. 10576-10583 ◽  
Author(s):  
Meddy El Alaoui ◽  
Alexandre Noiriel ◽  
Laurent Soulère ◽  
Lucie Grand ◽  
Yves Queneau ◽  
...  
Keyword(s):  
High Throughput ◽  
Phospholipase A ◽  
Microtiter Plates ◽  
High Throughput Assay

scholarly journals Development of a High-Throughput Screening Assay to Identify Inhibitors of the SARS-CoV-2 Guanine-N7-Methyltransferase Using RapidFire Mass Spectrometry

2021 ◽  
pp. 247255522110006
Author(s):  
Lesley-Anne Pearson ◽  
Charlotte J. Green ◽  
De Lin ◽  
Alain-Pierre Petit ◽  
David W. Gray ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Mutational Analysis ◽  
Nonstructural Protein ◽  
Translation Efficiency ◽  
Screening Assay ◽  
High Throughput Screening Assay ◽  
Starting Point ◽  
Approved Drugs ◽  
High Throughput Assay

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) represents a significant threat to human health. Despite its similarity to related coronaviruses, there are currently no specific treatments for COVID-19 infection, and therefore there is an urgent need to develop therapies for this and future coronavirus outbreaks. Formation of the cap at the 5′ end of viral RNA has been shown to help coronaviruses evade host defenses. Nonstructural protein 14 (nsp14) is responsible for N7-methylation of the cap guanosine in coronaviruses. This enzyme is highly conserved among coronaviruses and is a bifunctional protein with both N7-methyltransferase and 3′-5′ exonuclease activities that distinguish nsp14 from its human equivalent. Mutational analysis of SARS-CoV nsp14 highlighted its role in viral replication and translation efficiency of the viral genome. In this paper, we describe the characterization and development of a high-throughput assay for nsp14 utilizing RapidFire technology. The assay has been used to screen a library of 1771 Food and Drug Administration (FDA)-approved drugs. From this, we have validated nitazoxanide as a selective inhibitor of the methyltransferase activity of nsp14. Although modestly active, this compound could serve as a starting point for further optimization.

scholarly journals High-Throughput Peptide Derivatization toward Supramolecular Diversification in Microtiter Plates

ACS Nano ◽  
2021 ◽  
Author(s):  
Yiyang Lin ◽  
Matthew Penna ◽  
Christopher D. Spicer ◽  
Stuart G. Higgins ◽  
Amy Gelmi ◽  
...  
Keyword(s):  
High Throughput ◽  
Microtiter Plates ◽  
Peptide Derivatization

scholarly journals Development of a Novel Label-Free and High-Throughput Arginase-1 Assay Using Self-Assembled Monolayer Desorption Ionization Mass Spectrometry

2021 ◽  
pp. 247255522110006
Author(s):  
Michael D. Scholle ◽  
Zachary A. Gurard-Levin
Keyword(s):  
Mass Spectrometry ◽  
Drug Discovery ◽  
High Throughput ◽  
Ionization Mass Spectrometry ◽  
Ionization Mass ◽  
Label Free ◽  
Desorption Ionization ◽  
Arginase 1 ◽  
Self Assembled ◽  
High Throughput Assay

Arginase-1, an enzyme that catalyzes the reaction of L-arginine to L-ornithine, is implicated in the tumor immune response and represents an interesting therapeutic target in immuno-oncology. Initiating arginase drug discovery efforts remains a challenge due to a lack of suitable high-throughput assay methodologies. This report describes the combination of self-assembled monolayers and matrix-assisted laser desorption ionization mass spectrometry to enable the first label-free and high-throughput assay for arginase activity. The assay was optimized for kinetically balanced conditions and miniaturized, while achieving a robust assay (Z-factor > 0.8) and a significant assay window [signal-to-background ratio > 20] relative to fluorescent approaches. To validate the assay, the inhibition of the reference compound nor-NOHA (Nω-hydroxy-nor-L-arginine) was evaluated, and the IC50 measured to be in line with reported results (IC50 = 180 nM). The assay was then used to complete a screen of 175,000 compounds, demonstrating the high-throughput capacity of the approach. The label-free format also eliminates opportunities for false-positive results due to interference from library compounds and optical readouts. The assay methodology described here enables new opportunities for drug discovery for arginase and, due to the assay flexibility, can be more broadly applicable for measuring other amino acid–metabolizing enzymes.

A high-throughput assay for HIV-1 integrase 3′-processing activity using time-resolved fluorescence

2012 ◽  
Vol 184 (1-2) ◽  
pp. 34-40 ◽  
Author(s):  
Ying-Shan Han ◽  
Peter Quashie ◽  
Thibault Mesplede ◽  
Hongtao Xu ◽  
Kevork Mekhssian ◽  
...  
Keyword(s):  
High Throughput ◽  
Time Resolved Fluorescence ◽  
Time Resolved ◽  
Processing Activity ◽  
High Throughput Assay ◽  
Hiv 1
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