PUBLICATIONS

featuring the Echo acoustic technology

109 Total Publications

TITLES and AUTHORS

  • Year
  • Link
  • PDF
  • + Abstract
  • Optimizing the Profiling of Protein Kinases
  • Dimond P
  • Institution: Life Science Consultant
  • Publication: Genetic Engineering and Biotechnologies News, Vol 28, No. 10 (May 15, 2008)
  • 2008
  •  

Protein kinases comprise the largest family of mammalian enzymes, totaling at least 500. These ubiquitous enzymes play key roles in cellular regulation by catalyzing the reversible phosphorylation of more than 10,000 proteins.

Dysfunctional intracellular signaling through protein kinases is associated with about 400 human diseases, most notably cancers. As a result of the clinical success of the p210 Bcr-Abl inhibitor (Novartis’ Gleevec) for treatment of chronic myelogenous leukemia, pharma companies have applied massive resources to finding the next blockbuster kinase activity-modulating drug. And, as kinase-focused drug discovery enterprises expand, so does the need for discovery tools that automate and scale them.

At the Society for Biomolecular Screening’s (SBS) annual meeting held last month, companies collaborated in workshops, combining their technologies to address formidable challenges in kinase profiling and to facilitate selection of appropriate assay options.

In a workshop entitled “Assay Optimization and Kinase Profiling in micro HTS Format,” Labcyte (www.labcyte.com), Deerac Fluidics (www.deerac.com), Promega (www.promega.com), Corning Life Sciences (www.corning.com), and BMG Labtech (www.bmglabtech.com) teamed up to present a complete platform approach for kinase-assay development.

Using Labcyte’s ultralow volume-test technology, Corning Life Sciences’ specially developed dual assay plates, luminescent kinase-assay technology from Promega, Deerac Fluidics’ reagent-dispensing technology, and BMG Labtech’s PHERAstar luminescent-detection instrumentation, these companies demonstrated the complementarity and adaptability of their diverse technologies for the development of extremely low-volume, protein-kinase assays.

  • Assay Sciences: A Model for Improving Efficiency Through Centralization
  • Wingfield J, Jones D, Clark R, Simpson P
  • Institution: AstraZeneca
  • Publication:
  • 2007
  •  

The pharmaceutical industry is facing increasing external market pressures to reduce costs and deliver the next generation of therapeutics cost effectively. Pharmaceutical R&D faces the challenge of streamlining all stages of the drug discovery process. Delivering high quality decision-making data at the earliest opportunity serves to accelerate research and maintain product pipelines that are critical to business success. Reducing the time between Hit to Lead, whilst operating in an increasingly challenging business environment requires a different approach when deploying resources.

Secondary screening in the pharmaceutical industry has traditionally been carried out within project teams, in many cases biochemical and cell screening are carried out by the same bioscience resource. This typically results in a serial approach to screening where biochemical data is gathered before or during the development of any cellular assays. In a post high throughput (HTS) environment the level of automation applied to support screening has often been quite limited as there is no perceived benefit of automation when screening smaller numbers of samples. This model has the potential to delay project screening once the primary screening campaign has been completed, partly due to a hand over phase and repeat validation when the assay is handed back to project bioscientists. In some cases, screening may need to be suspended while adequate bioscience resource is redeployed from other on-going project support work. Moving away from the traditional project-centric lead identification to a centralised capability offers many advantages to both the bioscience and chemistry communities. Centralisation of both biochemical and cellular secondary screening activities enables flexible use of resources, allowing rapid responses to changing business demands.

Centralisation permits testing across multiple targets and increases the speed that data is generated. In addition focusing a small team on one aspect of drug discovery enables them to standardise the automation, technology, process and adopt best practices. This can result in a flexible, agile work team that can respond effectively to changing demand. Focusing teams in this way provides clarity around their expected delivery, which helps individuals maintain drive. On the counter side, there is a need to manage the perceived loss of project ownership of the assays that are centralised. The centralised team need to understand that their output will be subject to additional scrutiny. Initially there will need to be a considerable investment of time in managing customer interactions during the establishment of the capability, building customer confidence. Having a clear service level agreement (SLA) between the central screening team and each customer, which specifies the turn round time for data delivery, stringent QC process for data analysis, roles and responsibilities for individuals is important. The screening team then have to deliver against these SLA's.

  • Related Subject:high throughput, drug discovery
  • A High-Content Glucocorticoid Receptor Translocation Assay for Compound Mechanism-of-Action Evaluation
  • Agler M, Prack M, Zhu Y, Kolb J, Nowak K, Ryseck R, Shen D, Cvijic M, Somerville J, Nadler S, Chen T
  • Institution: Bristol-Myers Squibb, St. Jude Children's Hospital
  • Publication: J Biomol Screen December 207 vol. 12 no. 8 1029-1041, 10.1177/1087057107309353
  • 2007
  •  

Ligand-induced cytoplasm to nucleus translocation is a critical event in the nuclear receptor (NR) signal transduction cascade. The development of green fluorescent proteins and their color variants fused with NRs, along with the recent developments in automated cellular imaging technologies, has provided unique tools to monitor and quantify the NR translocation events. These technology developments have important implications in the mechanistic evaluation of NR signaling and provide a powerful tool for drug discovery. The unique challenges for developing a robust NR translocation assay include cytotoxicity accompanied with chronic overexpression of NRs, basal translocation induced by serum present in culture medium, and interference from endogenous NRs, as well as subcellular dynamics. The authors have developed a robust assay system for the glucocorticoid receptor (GR) that was applied to a panel of nuclear receptor ligands. Using a high-content imaging system, ligand-induced, dose-dependent GR nuclear translocation was quantified and a correlation with other conventional assays established.

  • Publication / Type:J Biomol Screen December 207 vol. 12 no. 8 1029-1041, 10.1177/1087057107309353
  • Related Subject:glucocorticoid receptor, nuclear translocation, high content screening, green fluorescent proteins, androgen receptor, mechanism of action
  • Link:http://jbx.sagepub.com/content/12/8/1029.full.pdf+html
  • Keeping DMSO Concentration Below 0.5% to Minimize its Effect in HTS Assays
  • Gallagher A, Koechlein D, Shieh J
  • Institution: Deerac Fluidics, Labcyte Inc.
  • Publication: J Biomol Screen December 207 vol. 12 no. 8 1029-1041, 10.1177/1087057107309353
  • 2007
  •  

Dimethyl sulfoxide (DMSO) is a commonly used solvent for compounds. DMSO accelerates protein unfolding and weakens the binding between small molecules and proteins. Consequently researchers keep DMSO concentrations as low as possible, especially for sensitive assays. To keep the DMSO concentration at less than one percent of the final assay volume has been difficult due to the lack of reliable nanoliter-range liquid handlers. Intermediate aqueous dilution steps can cause the compound to “crash out” of solution. The requirement to keep compounds dissolved while keeping DMSO concentration low in the final assay is especially critical when preparing compound activity curves. The Labcyte Echo™ 550 liquid handler utilizes acoustic drop ejection (ADE) to transfer 2.5-250 nL of compounds in DMSO directly from storage plates to assay plates. Deerac Fluidics Latitude™ bulk dispenser, which uses “spot-on” technology, can precisely deliver as low as 50 nL. The Latitude can be used to rapidly add pure DMSO to specific wells so that all assay wells have the same DMSO concentration. Here we demonstrate the use of these two technologies in combination for keeping final DMSO concentration under 0.5% in HTS assays.

  • Publication / Type:J Biomol Screen December 207 vol. 12 no. 8 1029-1041, 10.1177/1087057107309353
  • Related Subject:DMSO Concentration Below 0.5% Minimize Effect HTS Assays
  • Debunking the Myth: Validation of Fluorescein for Testing the Precision of Nanoliter Dispensing
  • Harris D, Mutz M
  • Institution: Labcyte Inc., Institute for Molecular Medicine Finland, FIMM, University of Helsinki
  • Publication: (JALA 2006;11:233–9) DOI: 10.1016/j.jala.2006.05.006
  • 2006
  •  

Acoustic energy can precisely and accurately eject a droplet of liquid from a reservoir, enabling delivery of picoliter and nanoliter volumes. Acoustic droplet ejection has been shown to be extremely precise (coefficients of variation !2%) over a wide range of dispensed volumes. However, measuring the performance of low-volume fluid transfers can be difficult because the data are often masked by variability in bulk dispensers and fluorescence readers used as part of the overall measurement process. The fluorophore used must also be stable so that thermal bleaching and photobleaching do not contribute additional variability to the measurements. This study assesses the suitability of fluorescein to measure the precision of fluid transfers of 2.5-nL DMSO droplets. The short-term and long-term stabilities of fluorescein are first qualified using a reference standard. Next, we determine the noise contribution of the filler and reader. Lastly, data are presented for the precision of 5- and 50-nL fluid transfers using this fluorescein measurement process.

  • The Effects of Small Molecule and Protein Solutes on Acoustic Drop Ejection
  • Shieh J, Ellson R, Olechno J
  • Institution: Labcyte Inc.
  • Publication: Journal of Laboratory AutomationAugust 2006 vol. 11 no. 4 227-232, 10.1016/j.jala.2006.06.005
  • 2006
  •  

Increasing the solute concentration of a solution alters the speed of sound in the solution in addition to changing the fluid viscosity and surface tension. For liquid handling devices that transfer fluids with acoustic energy, the change in sound speed can impact the focus of the acoustic energy and ultimately the accuracy of the transfer volume. To determine the effects of increasing solute concentration on transfer accuracy and precision, we studied the change in sound speed in dimethyl sulfoxide with the addition of low-molecular weight compounds and in phosphate-buffered saline with protein. The impact of compound and protein concentration on transfer precision and accuracy was found to be minimal and ADE can be used to transfer fluids regardless of these additions.

  • Improving IC50 Results with Acoustic Droplet Ejection
  • Olechno J, Shieh J, Ellson R
  • Institution: Labcyte Inc.
  • Publication: Journal of Laboratory Automation August 2006 vol 11 no. 4 240-246, 10.1016/j.jala.2006.06.007
  • 2006
  •  

IC50 analyses are typically sample, time, and labor intensive. They commonly require multiple dilution steps and consume significant amounts of sample compound. Aqueous intermediate dilutions of concentrated stock solutions can lead to rapid sample precipitation and the generation of false negatives. Hydrophobic compounds may stick to pipette tips or intermediate dilution vessels, reducing the concentration of the analyte in the dilution and also increasing the possibility of cross contamination. The requirement for multiple serial dilutions in common IC50 analyses causes significant accumulated error. Concentrations of dimethyl sulfoxide (DMSO), the typical solvent used to solubilize compound libraries, as low as 1% in the final assay solution can significantly affect the results of the experiment. Finally, the cost of pipette tips and intermediate dilution vessels, and the frequency of the DMSO washes of tips grows significantly as the number of compounds being analyzed is increased. A system incorporating acoustic droplet ejection of compounds improves IC50 results by reducing the amount of sample used in the analysis to nanoliters, eliminating intermediate aqueous dilutions and accumulated pipetting error, lowering DMSO concentrations in the final assay to below 1%, and reducing costs of consumables (plastics, solvents, and their disposal).

  • Publication / Type:Journal of Laboratory Automation August 2006 vol 11 no. 4 240-246, 10.1016/j.jala.2006.06.007
  • Related Subject:ADE, DMSO impact on assays, aqueous intermediate dilutions, dose-response, Echo, dose-response, IC50, cost reduction, throughput, cross-contamination
  • Link:http://jla.sagepub.com/content/11/4/240.full.pdf+html
  • In situ DMSO Hydration Measurements of HTS Compound Libraries
  • Ellson R, Stearns R, Mutz M, Brown C, Browning B, Harris D, Qureshi S, Shieh J and Wold D.
  • Institution: Labcyte Inc.
  • Publication: Combinatorial Chemistry & High Throughput Screening, 8(6):489-498, 2005, DOI: 10.2174/13862070548673
  • 2005
  •  

Compounds used in high throughput screening (HTS) are typically dissolved in DMSO. These solutions are stored automation-friendly racks of wells or tubes. DMSO is hygroscopic and quickly absorbs water from the atmosphere. When present in DMSO compound solutions, water can accelerate degradation and precipitation. Understanding DMSO hydration in an HTS compound library can improve storage and screening methods by managing the impact of water on compound stability. A non-destructive, acoustic method compatible with HTS has been developed to measure water content in DMSO solutions. Performance of this acoustic method was compared with an optical technique and found to be in good agreement. The accuracy and precision of acoustic measurements was shown to be under 3% over the tested range of DMSO solutions (0% to 35% water by volume) and insensitive to the presence of HTS compounds at typical storage concentrations. Time course studies of hydration for wells in 384-well and 1536-well microplates were performed. Well geometry, fluid volume, well position and atmospheric conditions were all factors in hydration rate. High rates of hydration were seen in lower-volume fills, higher-density multi-well plates and when there was a large differential between the humidity of the lab and the water content of the DMSO. For example, a 1536-well microplate filled with 2μL of 100% DMSO exposed for one hour to a laboratory environment with ∼40% relative humidity will absorb over 6% water by volume. Understanding DMSO hydration rates as well as the ability to reverse library hydration are important steps towards managing stability and availability of compound libraries.

  • Publication / Type:Combinatorial Chemistry & High Throughput Screening, 8(6):489-498, 2005, DOI: 10.2174/13862070548673
  • Related Subject:DMSO hydration, acoustic hydration measurement, quality control, microtiter plates
  • Link:http://www.eurekaselect.com/60394/article
  • Transfer of low nanoliter volumes between microplates using focused acoustics—automation considerations
  • Ellson R, Mutz M, Browning B, Lee L, Miller M and Papen R
  • Institution: Labcyte Inc.and BMG Labtech
  • Publication: Journal of the Association for Laboratory Automation (Impact Factor: 1.5). 01/2003; 8(5):29-34. DOI:
  • 2003
  •  

Acoustic droplet ejection (ADE) gently and precisely aliquots nanoliter and picoliter liquid volumes without any physical contact with the solution being transferred. The technology is very automation-friendly, as it is compatible with conventional microplates. Focused energy from an acoustic transducer induces droplet ejection into an inverted standard microplate. The commercial system transfers low-nanoliter volumes of dimethyl sulfoxide–dissolved compound libraries and thereby enables cell-based assays to be performed in 1536-well plates.