Development of a Platform to Enable Fully Automated Cross-Titration Experiments. (April 2017)

• Record Type: Journal Article
• Authors: Cassaday, Jason; Finley, Michael; Squadroni, Brian; Jezequel-Sur, Sylvie; Rauch, Albert; Gajera, Bharti; Uebele, Victor; Hermes, Jeffrey; Zuck, Paul
• Is Part Of: SLAS technology. Volume 22:Number 2(2017)
• Page Start: 195
• Page End: 205
• Publication Date: 2017-04
• Subjects: agonist shift -- allostery -- automated biology -- engineering -- G protein–coupled receptor -- HTS -- high-throughput screening -- robotics and instrumentation; Medical laboratory technology -- Periodicals; Laboratories -- Equipment and supplies -- Periodicals; Diagnosis, Laboratory -- Periodicals; 616.075
• DOI: 10.1177/2211068216679805
• Languages: English
• ISSNs: 2472-6303

Abstract

In the triage of hits from a high-throughput screening campaign or during the optimization of a lead compound, it is relatively routine to test compounds at multiple concentrations to determine potency and maximal effect. Additional follow-up experiments, such as agonist shift, can be quite valuable in ascertaining compound mechanism of action (MOA). However, these experiments require cross-titration of a test compound with the activating ligand of the receptor requiring 100–200 data points, severely limiting the number tested in MOA assays in a screening triage. We describe a process to enhance the throughput of such cross-titration experiments through the integration of Hewlett Packard's D300 digital dispenser onto one of our robotics platforms to enable on-the-fly cross-titration of compounds in a 1536-well plate format. The process handles all the compound management and data tracking, as well as the biological assay. The process relies heavily on in-house-built software and hardware, and uses our proprietary control software for the platform. Using this system, we were able to automate the cross-titration of compounds for both positive and negative allosteric modulators of two different G protein–coupled receptors (GPCRs) using two distinct assay detection formats, IP1 and Ca 2+ detection, on nearly 100 compounds for each target.