A flow cytometric assay for simultaneous assessment of drug efflux, proliferation, and apoptosis.
BACKGROUND
Proliferative status and multidrug resistance play a key role in determining cell response to chemotherapy. There is a need to develop multiple labeling method that allows simultaneous assessment of multidrug resistant (MDR) phenotype, proliferative status, apoptosis related changes in mitochondrial potential, in chemosensitive and chemoresistant tumor cell populations.


METHODS
A three-color labeling was performed using Hoechst 33342 (DNA), JC1 (mitochondrial potential), and a far red fluorescent membrane intercalating dye: PTIR271 (proliferation).


RESULTS
Combined staining of DNA and mitochondrial potential allows identification of subpopulations expressing and MDR phenotype mediated by P-glycoprotein (Pgp), and, in Pgp negative subpopulations, identification of apoptotic cells and evaluation of cell cycle status in viable cells. Addition of a far red fluorescent membrane intercalating dye, PTIR271, allows simultaneous monitoring of cell division status by dye dilution in both drug sensitive and drug resistant populations.


CONCLUSIONS
This triple labeling is an interesting method to study the proliferation status of drug sensitivity and drug resistance in viable tumor cells.
Barbier
M
M
Laboratoire de Dynamique Cellulaire, E.P.H.E, UMR CNRS 5525, Institut Albert Bonniot, La Tronche, France.
Gray
B D
BD
Muirhead
K A
KA
Ronot
X
X
Boutonnat
J
J
D000972:Antineoplastic Agents, Phytogenic / Q000493:pharmacokinetics* / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000493:pharmacokinetics* / Q000494:pharmacology*; D002455:Cell Division / Q000187:drug effects; D019008:Drug Resistance, Neoplasm; D005434:Flow Cytometry / Q000379:methods*; D005456:Fluorescent Dyes; D006801:Humans; D000077146:Irinotecan; D020014:K562 Cells; D011237:Predictive Value of Tests; D013194:Staining and Labeling / Q000379:methods*
D000972:Antineoplastic Agents, Phytogenic; D005456:Fluorescent Dyes; D000077146:Irinotecan; D002166:Camptothecin