Dichloro-dihydro-fluorescein diacetate (DCFH-DA) assay: A quantitative method for oxidative stress assessment of nanoparticle-treated cells

被引:406
作者
Aranda, A. [1 ]
Sequedo, L. [1 ]
Tolosa, L. [1 ]
Quintas, G. [2 ]
Burello, E. [4 ]
Castell, J. V. [1 ]
Gombau, L. [3 ]
机构
[1] Hosp Univ Fe, Res Ctr, Unit Expt Hepatol, Valencia 46009, Spain
[2] Hosp Univ Fe, Res Ctr, LEITAT Technol Ctr, Analyt Unit, Valencia 46009, Spain
[3] LEITAT Technol Ctr, BioInvitro Div, Barcelona 08028, Spain
[4] Commiss European Communities, Joint Res Ctr, Inst Hlth & Consumer Protect, Syst Toxicol Unit, I-21027 Ispra, Italy
关键词
DCFH-DA; Hepatocytes; In vitro assays; Nanoparticles; CARBON NANOTUBES; CYTOTOXICITY; TOXICITY; AGGREGATION;
D O I
10.1016/j.tiv.2013.01.016
中图分类号
R99 [毒物学(毒理学)];
学科分类号
100405 ;
摘要
No consensus exists on how to address possible toxicity of nanomaterials as they interfere with most in vitro screening tests based on colorimetric and fluorimetric probes such as the dichloro-dihydrofluorescein diacetate (DCFH-DA) assay for detection of oxidative species. In the present research, nanomaterial interaction with DCFH-DA was studied in relation to its nature and/or assay conditions (cell-based and time exposure) by incubating Rhodamine (Rhd)-labeled 25 nm and 50 nm silica (SiO2), naked and oleic acid coated magnetite, (Fe3O4) and maghemite (Fe2O3) iron oxide, titanium dioxide (TiO2) and poly(ethylene oxide)-poly(lactide/glycolide) acid (PLGA-PEO) nanoparticles (NPs) with metabolically active rat hepatocytes for 4 and 24-h periods. Data indicated that nanoparticle uptake correlated with quenching of dye fluorescence emission. In spite of their masking effect, the oxidative potential of NPs could be detected at a limited threshold concentration when exposed for periods of time longer than those frequently used for this test. However, changes in the experimental conditions did not systematically result in free radical formation for all nanomaterials tested. Overall data indicate that despite the quenching effect of nanoparticles on DCFH-DA assay, it can be considered as a useful tool for quantitative measurement of NPs-induced oxidative stress by minor modifications of standardized protocols. (C) 2013 Elsevier Ltd. All rights reserved.
引用
收藏
页码:954 / 963
页数:10
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