title: Feasibility evaluation of 3 automated cellular drug screening assays on a robotic workstation.
abstract: This study presents the implementation and optimization of 3 cell-based assays on a TECAN Genesis workstation-the Caspase-Glo 3/7 and sulforhodamine B (SRB) screening assays and the mechanistic Caco-2 permeability protocol-and evaluates their feasibility for automation. During implementation, the dispensing speed to add drug solutions and fixative trichloroacetic acid and the aspiration speed to remove the supernatant immediately after fixation were optimized. Decontamination steps for cleaning the tips and pipetting tubing were also added. The automated Caspase-Glo 3/7 screen was successfully optimized with Caco-2 cells (Z' 0.7, signal-to-base ratio [S/B] 1.7) but not with DU-145 cells. In contrast, the automated SRB screen was successfully optimized with the DU-145 cells (Z' 0.8, S/B 2.4) but not with the Caco-2 cells (Z' -0.8, S/B 1.4). The automated bidirectional Caco-2 permeability experiments separated successfully low- and high-permeability compounds (Z' 0.8, S/B 84.2) and passive drug permeation from efflux-mediated transport (Z' 0.5, S/B 8.6). Of the assays, the homogeneous Caspase-Glo 3/7 assay benefits the most from automation, but also the heterogeneous SRB assay and Caco-2 permeability experiments gain advantages from automation.
authors: Division of Pharmaceutical Technology, Faculty of Pharmacy, University of Helsinki, Finland. anne.soikkeli@helsinki.fi
authors: Anne
authors: A
authors: Soikkeli
authors: Cristina
authors: C
authors: Sempio
authors: Ann Marie
authors: AM
authors: Kaukonen
authors: Arto
authors: A
authors: Urtti
authors: Jouni
authors: J
authors: Hirvonen
authors: Marjo
authors: M
authors: Yliperttula
mesh_terms: D001331:Automation; D001681:Biological Assay / Q000295:instrumentation* / Q000379:methods*; D018938:Caco-2 Cells; D053148:Caspase 3 / Q000378:metabolism; D053179:Caspase 7 / Q000378:metabolism; D002463:Cell Membrane Permeability; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation* / Q000379:methods*; D005240:Feasibility Studies; D006801:Humans; D012235:Rhodamines / Q000378:metabolism; D012371:Robotics / Q000295:instrumentation*
chemical_list: D012235:Rhodamines; C022027:lissamine rhodamine B; D053148:Caspase 3; D053179:Caspase 7