Parameters influencing the transport rate of marker compounds across Caco-2 monolayers

Performing transport experiments with Caco-2 monolayers some important parameters have to be considered. Cell layers are appropriate for transport studies after complete polarisation and differentiation. Consequently they should be used 20 - 25d after seeding. Monolayers grown for shorter time than 20d yield to high transport rates, a cultivation period longer than 25d results in layers susceptible to detachment. Additionally the material of the filter support exerts an influence on the transport data because of varying barrier functions of cell layers grown on polyester- versus polycarbonate filter inserts. By comparison of transport rates obtained with different model systems such as Transwells and Ussing chambers the influence of the unstirred water layer on permeation of markers was investigated. The results indicate that there is a noticeable difference with hydrophobic compounds only. To check the dependence of transport phenomena on energy the difference in transport rates between 4°C and 37°C as well as the cumulative transport subject to the transport direction was determined. Moreover the influence of the stirring rate on the cumulative transport in the Ussing chamber system was elucidated. For assessment of the cell associated amount of the transported substance the cell layer was dissolved in NaOH / SDS to get further valuable informations.