Studies on Repository Compound Stability in DMSO under Various Conditions

Cheng, Xueheng; Hochlowski, Jill E.; Tang, Hua; Hepp, Darlene; Beckner, Carl F.; Kantor, Stan; Schmitt, Robert J.

doi:10.1177/1087057103008003007

Cited by 101 publications

(85 citation statements)

References 8 publications

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“…Given that both assay reagents and compounds in the screening collection may exhibit temporal variations in activity [30,31], we optimized the screening protocol to test each compound sample in the two individual fluorophore assays as closely in time as possible. There was no specific way within the robotic software to ensure that assay plates were run in an alternating fashion between the green and red systems, or set up any explicit dependencies or contingencies between the two assays.…”

Section: Interleaved Dual-assay Qhtsmentioning

confidence: 99%

Dual-fluorophore quantitative high-throughput screen for inhibitors of BRCT–phosphoprotein interaction

Simeonov

Yasgar

Jadhav

et al. 2008

Analytical Biochemistry

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Finding specific small-molecule inhibitors of protein-protein interactions remains a significant challenge. Recently, attention has grown toward "hot-spot" interactions where binding is dominated by a limited number of amino acid contacts, theoretically offering an increased opportunity for disruption by small molecules. Inhibitors of the interaction between BRCT (Cterminal portion of BRCA1, a key tumor suppressor protein with various functions), and phosphorylated protein (Abraxas, BACH1, CtIP) implicated in DNA damage response and repair pathways, should prove useful in studies of BRCA1's role in cancer and to potentially sensitize tumors to chemotherapeutic agents. We developed and miniaturized to 1536-well format and 3 μL final volume a pair of fluorescence polarization (FP) assays utilizing fluorescein-and rhodaminelabeled pBACH1 fragment. In order to minimize the effect of fluorescence artifacts and to increase the overall robustness of the screen, the 75,552 compound library members were each assayed against both the fluorescein-and rhodamine-labeled probe-protein complexes in separate but interleaved reactions. In addition, every library compound was tested over a range of concentrations, following the qHTS paradigm (Inglese et al, PNAS, 103, 1147(2006). Analyses of the screening results led to the selection and subsequent confirmation of 16 compounds active in both assays. Faced with a traditionally difficult protein-protein interaction assay, by performing two-fluorophore qHTS we were able to confidently select a number of actives for further studies.

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Section: Interleaved Dual-assay Qhtsmentioning

confidence: 99%

Dual-fluorophore quantitative high-throughput screen for inhibitors of BRCT–phosphoprotein interaction

Simeonov

Yasgar

Jadhav

et al. 2008

Analytical Biochemistry

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“…30 DMSO is hygroscopic and rapidly absorbs water from the atmosphere, and studies have shown that water uptake in compound DMSO solutions significantly contributes to compound degradation, precipitation, and reduction in concentration. 31,32 An open Echo source plate filled with 100% DMSO at under half capacity will equilibrate to a moisture content of over 30% within 24 h under standard laboratory conditions 23 ; therefore, storage conditions aimed at limiting hydration both prior to and during use are an important consideration. In AP, compound source plates for cell assays were created up to 1 week before the assay, foil sealed, and stored within a StoragePod nitrogen enclosure at 5% relative humidity (Roylan Developments Ltd., Leatherhead, Surrey, UK).…”

Section: Integrated Echo System I-postimplementation Enhancementsmentioning

confidence: 99%

Implementation and Challenges of Direct Acoustic Dosing into Cell-Based Assays

et al. 2016

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Since the adoption of Labcyte Echo Acoustic Droplet Ejection (ADE) technology by AstraZeneca in 2005, ADE has become the preferred method for compound dosing into both biochemical and cell-based assays across AstraZeneca research and development globally. The initial implementation of Echos and the direct dosing workflow provided AstraZeneca with a unique set of challenges. In this article, we outline how direct Echo dosing has evolved over the past decade in AstraZeneca. We describe the practical challenges of applying ADE technology to 96-well, 384-well, and 1536-well assays and how AstraZeneca developed and applied software and robotic solutions to generate fully automated and effective cell-based assay workflows.

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“…Previous studies have typically characterized the stability of compounds in a lowvolume (<40 µL) 96-or 384-well plate format, which differ significantly from the high-volume (1.4-mL) Matrix tubes that Amgen has standardized for archive compound storage. 4,[6][7][8] As we will demonstrate, the labware format can affect the experimental parameters, so it is important to consider the storage vessel form factor in any stability validation study. We also wanted to characterize the effectiveness of a resolubilization protocol using a Covaris AFA platform.…”

mentioning

confidence: 99%

Characterization of Short-Term Temperature, Exposure, and Solubilization Effects on Library Compound Quality

et al. 2011

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IntroductIon Multiple groups have recognized that maintaining compound quality is dependent on both storage conditions and sample handling methods.1-3 Our workflows within Amgen have been built around low-volume, single-use operational tubes that are periodically prepared as replicates from high-volume, multiuse archive tubes, both stored at temperatures of -8 or -20 °C. This design minimizes the number of times an archive tube must be accessed by instead relying on the single-use tubes for normal operations. This approach effectively serves the majority of the collection, but we have recently noticed that certain compounds enter stages of high demand for a shortened period (<3 months) of time. These high-demand compounds often require frequent replenishment from multiuse archive tubes, which can affect the number of freeze-thaw cycles a compound undergoes. The extended sample handling time can lead to a decrease in overall order fulfillment efficiency and degradation in sample quality from unnecessary exposure outside of the store. Although one potential solution might be to prepare more initial single-use tubes, it is often hard to predict how much material will be required, and the preferred approach has been to replenish on-demand.One alternative approach is short-term storage at room temperature, which would eliminate the need for thawing and reduce sample handling time. Other groups have shown that compound quality can remain stable for 4 to 6 months 4-6 at room temperature, but the labware was significantly different from our archive operations, leading to questions around the form factor used to store compounds. In this study, we wanted to understand the impact of short-term storage at room temperature as compared to the effect of multiple freeze-thaw cycles on overall compound quality in labware relevant to our operations. For our characterization of freeze-thaw events, we were primarily interested in the cumulative effect of both the freeze-thaw and the water uptake that is common with sample handling during formatting Many compound collections are stored under the same temperature conditions, which can limit flexibility by increasing the processing time required for high-demand compounds. In this study, the authors wanted to evaluate the impact of a hybridstorage approach where high-demand compounds are stored for a shortened time period at room temperature to expedite processing operations. The use of a Covaris adaptive-focused acoustics platform was also characterized as a potential enhancement or alternative to storage at elevated temperatures. This study evaluated the impact of temperature, exposure, and solubilization on overall compound quality for short-term storage. A small library of 25 representative compounds was evaluated over an 18-week period to monitor the change in purity and concentration by high-performance liquid chromatography with ultraviolet detection. The authors concluded that temperature had a significant impact on compound concentration, and the effects due to exposure cycles...

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Studies on Repository Compound Stability in DMSO under Various Conditions

Cited by 101 publications

References 8 publications

Dual-fluorophore quantitative high-throughput screen for inhibitors of BRCT–phosphoprotein interaction

Dual-fluorophore quantitative high-throughput screen for inhibitors of BRCT–phosphoprotein interaction

Implementation and Challenges of Direct Acoustic Dosing into Cell-Based Assays

Characterization of Short-Term Temperature, Exposure, and Solubilization Effects on Library Compound Quality

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