Permeability analysis - PAMPA

The Parallel Artificial Membrane Permeability Assay (PAMPA) is the non-cell based assay for predicting passive absorption of drugs in early drug discovery. It uses a lipid filled membrane to simulate the lipid bilayer of various cell types, including intestinal epithelium. Most drugs are absorbed through the intestines without using cellular pumps. Therefore, passive permeability assays are useful for screening oral-absorption potential of drug candidates. The PAMPA’s results correlate with human drug absorption value and its ability to mimic the target barrier can by enhanced by the optimization of the assay’s conditions.

Method: The PAMPA assay is based on the use of a 96-well membrane filter based plate. The aim of the method is to determine the ability of compounds to diffuse from a donor into an acceptor compartment separated by a 0.45 µm PVDF membrane filter pretreated with a lipid-containing organic solvent. The artificial membrane is built up by pipetting a solution of lipids on a supporting filter material in a donor plate which is then filled with buffer containing the tested compound. The plate is then placed on a 96-well acceptor plate filled with buffer solution to insure the contact of liquids. The plates are incubated together for 2-16 hours and after compound diffusion the donor plate is removed from the acceptor plate. The contents of the acceptor and donor wells are replaced to a UV compatible 96-well plate and concentrations of the compounds are analyzed in a UV/Vis microtiter plate reader.