Risorse - Poster scientifici
Use of a Flexible Multipurpose Automated 1536-well Microplate Dispenser for HTSDownload
September 08, 2010
Authors: Paul Held, Gary Prescott, Andreas Rieger, Jürg Wetterwald, Brad Larson, and Peter Banks; BioTek Instruments, Inc., Winooski, Vermont, USA
Today’s HTS demands require that automated dispensers are not only fast and accurate, but also are fl exible, compact and serve multiple tasks. Traditional screening technologies employed in vitro assays to screen potential drug compounds for activity. However, with advances in HTS screening, drug discovery has migrated toward cell-based assays performed in high-density microplates. The use of 1536-well microplates for cell-based assays requires the use of accurate and reliable automation in order to dispense uniform numbers of cells to each microplate well in a volume of a few microliters. As with any cell-based experiment, providing uniform numbers of viable cells is paramount to successful experiments. CHO-M1 cells (200 cells/μL) were dispensed into 1536-well microplates and cell uniformity as measured by the luminescent determination of ATP. Luminescent signal for wells not containing cells averaged 10, while those wells with 800 cells averaged 23210 with a Z’ =0.753.
In addition, unique 3-D microcarrier culturing systems such as GEM™ (Global Eukaryotic Microcarrier) have also been automated. These small sub-micron magnetic optically clear particles serve as a carrier for cells avoiding trypsinization prior to plate seeding. We demonstrate the ability of this 3-D cell culture method to measure the activity of the Histamine H1 G-protein-coupled receptor using a FRET-based assay. The entire assay procedure was automated in 384-well format.
The phospho-activation of ERK1/2 can serve as a surrogate marker for G-protein dependent and G-protein independent signals. However, until recently HTS-compatible technologies for measuring ERK activation have been lacking. We have automated a BacMam ERK1/2 Cellular Assay that combines BacMam-mediated gene delivery of a GFP-ERK2 sensor with LanthaScreen® Cellular Assay technology for the measurement of intracellular phospho-ERK1/2 levels. This methodology was used to measure agonist pERK1/2 response at high Z’- factor levels, as well as screen a Tocris library biased towards GPCR and ion channel binding compounds for D2 dopamine receptor antagonists in 384-well microplates.
Here we describe the performance of the MultiFlo dispenser to dispense a number of different reagents under various experimental conditions. Details regarding work fl ow and instrument performance will be provided as measured by biologically relevant responses in several different experiments.