TR146 CELLS AS A MODEL FOR HUMAN BUCCAL EPITHELIUM .2. OPTIMIZATION AND USE OF A CELLULAR-SENSITIVITY MTS PMS ASSAY/

The optimisation and use of the cellular sensitivity MTS/PMS assay for TR146 cells are presented. The MTS/PMS assay is a colorimetric method based on reduction of a tetrazolium salt to the corresponding formaza n including an electron acceptor as accelerator. The optimisation stud ies resulted in a final protocol for drug sensitivity testing of TR146 cells. The initial cell density was 2 x 10(4) cells/well, and after 4 h incubation with a MTS/PMS reagent (final concentrations: MTS, 240 m u g/ml; PMS, 2.4 mu g/ml), the optical density was recorded at 490 nm. The optical density of the TR146-generated MTS-formazan at room tempe rature and atmosphere was shown toe constant for a period of 30 min. T he final protocol for the MTS/PMS assay used for a series of beta-adre noceptor antagonists (propranolol, oxprenolol, metoprolol, and atenolo l) in concentrations in the range 10(-6)-10(-2) M indicated that solut ions of about 10(-4) M caused a 10% decrease of the cellularly generat ed MTS-formazan (i.e. IC10 = 10(-4) M). The assay used for a series of morphine and morphine prodrugs (3-hexanoyl-, 3-propionyl- and 3-acety l-morphine) in concentrations in the range 3.5 x 10(-7)-3.5 x 10(-3) M showed that only the most lipophilic substance, 3-hexanoyl-morphine, affected the TR146 cells (IC10= 10(-4) M).