CYTOFLUOROMETRIC DETERMINATION OF THE P-G LYCOPROTEIN-MEDIATED MULTIDRUG-RESISTANCE

Multidrug resistance (MDR) can be mediated by transmembrane glycoprote in P-170 serving as the energy-dependent efflux pump for diverse lipop hylic toxic compounds. A spectrofluorimetric method based on automatic or semi-automatic measurement of the fluorescence of cell-associated dyes, Rhodamine 123 and Hoechst 33258, allowed detection of MDR cells. Human malignancies often exhibit low levels of Pgp. Therefore it is r easonable to detect early stages of Pgp activity. A simple test for Pg p-mediated efflux was devised. The suitability of this technique for t he assessment of low-rate Pgp-mediated efflux in tumor cell lines and in human PDL (in CML patients) was studied. The method allowed us to d etermine cultured human tumour cells with low (about 2-fold) level of resistance to the selective agent colchicine and also to discriminate the minority (10%) of MDR cells in mixed cell populations. The data ob tained by staining cells with fluorescent dyes well correlated with th e results of an indirect immunofluorescence assessment of P-170 using monoclonal antibodies UIC-2.