Flow cytometric double labeling technique for screening of multidrug resistance.
We investigated the capabilities of flow cytometry in the analysis of a multidrug resistant (MDR) human ovarian cancer cell line 2780AD and its drug sensitive parental A2780. A functional assay using daunorubicin (DNR) as a fluorescent probe was combined with an immunofluorescence assay of P-glycoprotein (P-gp) using the monoclonal antibody MRK-16. Functionally MDR could be demonstrated by the lower DNR-content of MDR cells compared to DNR-content of drug sensitive cells. When incubation was performed with DNR in the presence of verapamil, DNR-content increased in the MDR cells. However the content of the A2780 cells was never attained. Differences in DNR-content were not related to differences in DNA-content. In experimental cell lines immunofluorescence data were inversely related with those of DNR-content: MDR cells had high levels of P-gp expression and low levels of DNR-content (and vice versa in drug sensitive cells). Both assays can be easily combined in a multiparametric flow cytometric procedure to evaluate both parameters simultaneously in the same cells. Analysis of clinical samples demonstrates the existence of aberrant subpopulations which would not be detected by using a single parameter assay.
Lab of Cancer Research and Clinical Oncology, Antwerp University, Wilrijk, Belgium.
E E
EE
Gheuens
D R
DR
van Bockstaele
M
M
van der Keur
H J
HJ
Tanke
A T
AT
van Oosterom
E A
EA
De Bruijn
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000903:Antibiotics, Antineoplastic / Q000032:analysis*; D000911:Antibodies, Monoclonal; D002461:Cell Line, Transformed; D003630:Daunorubicin; D004351:Drug Resistance; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D005456:Fluorescent Dyes; D006801:Humans; D008562:Membrane Glycoproteins / Q000032:analysis* / Q000378:metabolism; D010051:Ovarian Neoplasms / Q000378:metabolism
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000903:Antibiotics, Antineoplastic; D000911:Antibodies, Monoclonal; D005456:Fluorescent Dyes; D008562:Membrane Glycoproteins; D003630:Daunorubicin