title: Real-time cell viability assays using a new anthracycline derivative DRAQ7®.
abstract: The exclusion of charged fluorescent dyes by intact cells has become a well-established assay for determining viability of cells. In search for a noninvasive fluorescent probe capable of long-term monitoring of cell death in real-time, we evaluated a new anthracycline derivative DRAQ7. The novel probe does not penetrate the plasma membrane of living cells but when the membrane integrity is compromised, it enters and binds readily to nuclear DNA to report cell death. It proved to be nontoxic to a panel of cancer cell lines grown continuously for up to 72 h and did not induce any detectable DNA damage signaling when analyzed using laser scanning microscopy and flow cytometry. The DRAQ7 provided a sensitive, real-time readout of cell death induced by a variety of stressors such as hypoxia, starvation, and drug-induced cytotoxicity. The overall responses to anticancer agents and resulting pharmacological dose-response profiles were not affected by the growth of tumor cells in the presence DRAQ7. Moreover, we for the first time introduced a near real-time microflow cytometric assay based on combination of DRAQ7 and mitochondrial inner membrane potential (ΔΨ(m) ) sensitive probe TMRM. We provide evidence that this low-dosage, real-time labeling procedure provides multiparameter and kinetic fingerprint of anticancer drug action.
authors.affiliation: The BioMEMS Research Group, School of Chemical Sciences, University of Auckland, Auckland, New Zealand.
authors.forename: Jin
authors.initials: J
authors.lastname: Akagi
authors.forename: Magdalena
authors.initials: M
authors.lastname: Kordon
authors.forename: Hong
authors.initials: H
authors.lastname: Zhao
authors.forename: Anna
authors.initials: A
authors.lastname: Matuszek
authors.forename: Jurek
authors.initials: J
authors.lastname: Dobrucki
authors.forename: Rachel
authors.initials: R
authors.lastname: Errington
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Smith
authors.forename: Kazuo
authors.initials: K
authors.lastname: Takeda
authors.forename: Zbigniew
authors.initials: Z
authors.lastname: Darzynkiewicz
authors.forename: Donald
authors.initials: D
authors.lastname: Wlodkowic
mesh_terms: D018943:Anthracyclines / Q000737:chemistry* / Q000494:pharmacology; D017209:Apoptosis; D001549:Benzamides / Q000494:pharmacology; D001713:Biphenyl Compounds / Q000494:pharmacology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004249:DNA Damage; D003609:Dactinomycin / Q000494:pharmacology; D005047:Etoposide / Q000494:pharmacology; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000494:pharmacology; D006657:Histones / Q000378:metabolism; D006801:Humans; D020128:Inhibitory Concentration 50; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D009596:Nitrophenols / Q000494:pharmacology; D010879:Piperazines / Q000494:pharmacology; D012235:Rhodamines / Q000737:chemistry; D019311:Staurosporine / Q000494:pharmacology; D013449:Sulfonamides / Q000494:pharmacology; D013450:Sulfones / Q000494:pharmacology
chemical_list: C501332:ABT-737; D018943:Anthracyclines; D001549:Benzamides; D001713:Biphenyl Compounds; C582241:DRAQ7 fluorescent probe; D005456:Fluorescent Dyes; C468783:H2AX protein, human; D006657:Histones; D009596:Nitrophenols; D010879:Piperazines; D012235:Rhodamines; D013449:Sulfonamides; D013450:Sulfones; C515128:TW-37 compound; C401833:tetramethylrhodamine methyl ester; D003609:Dactinomycin; D005047:Etoposide; D019311:Staurosporine