P-glycoprotein mediated multidrug resistance assessment by flow-cytometry in malignant hemopathies

A relatively common and frequent form of multidrug resistance(MDR) in cancer cells is due to membrane overexpression of P-glycoprotein. Mdr phenotype was investigated by flow-cytometry in several types of malignant hemopathies -chronic lymphocytic leukemia, non-Hodgkin lymphomas, acute lymphoblastic and myeloblastic leukemias. We used daunomycin and fluo-3 as fluorochromes, and verapamil as reversor agent. The method is lacking unitary clinical parametrization and in order to improve it, we tried to establish an optimal concentration of verapamil, which was shown to be 14.92 micrograms/ml. The reliability of results obtained with fluo-3 in culture media containing Ca2+ is questionable, as low variations in the intracellular level of this ion dramatically influences light emission by the fluorochrome and possibly the function of P-gp. To avoid such fluorescence intensity variations, Ca(2+)-free cell culture medium for fluo-3-based flow-cytometric assay is suggested to be used.