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P2006
Automation of Pre-coated PAMPA Plates Improves Predictability, Reproducibility, & Efficiency
Presenter Kevin Chen, BD Biosciences, USA
Additional Authors: Xiaoxi (Kevin) Chen, Anthony Murawski, Michael Shanler, Linda Hladik, Charles L. Crespi
PAMPA (Parallel Artificial Membrane Permeability Assay) is a well-accepted screening assay for ADME properties (membrane permeability). It is in a 96-well plate format suitable for automation. Recently, BD Biosciences has developed a 96-well pre-coated PAMPA plate in which the microporous filters are pre-coated with a new artificial membrane – structured layers of phospholipids that mimic the biological membrane. The pre-coated PAMPA plate is stable for a prolonged period of time when stored at -20º C. The new artificial membrane produces correct prediction for the compounds under-predicted by traditional PAMPA. In this study we validated the pre-coated PAMPA plates using a set of compound standards.
The PAMPA experiments were carried out using a Tecan Genesis 150 liquid handler that is integrated to a plate reader using a robot arm. The overall time required to run the assay on the pre-coated plate is greatly reduced when compared to traditional methods, since effort is not required to form and prepare the membrane prior to adding the compounds. The instrument time is focused on dilution of compounds from a mother plate, addition of compounds to the Donor wells, incubation time, and then creation of the equilibrium plate and removal of samples for analysis from the Acceptor wells. The concentrations of compound solutions (acceptor solution & donor solution) were characterized using UV absorption. The automated method required fewer robotic steps than traditional automated PAMPA procedure, which resulted in improved process efficiency. Additionally, results are compared between automated and manual methods, showing good reproducibility compared to published literature values. This study demonstrates a convenient, sensitive and reproducible assay that allows rapid evaluation of passive drug permeability.