A micromethod for the in vitro micronucleus assay

Nesslany, Fabrice; Marzin, Daniel

doi:10.1093/mutage/14.4.403

Cited by 54 publications

(16 citation statements)

References 36 publications

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“…The in vitro micronucleus assay was performed according the procedure described by Nesslany and Marzin (1999). This micro-method used mouse lymphoma cells L5178Y.…”

Section: Aquatic (Geno)toxicity Testsmentioning

confidence: 99%

Comparison of solid and liquid-phase bioassays using ecoscores to assess contaminated soils

Lors

Ponge²,

Aldaya³

et al. 2011

Environmental Pollution

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Bioassays on aqueous and solid phases of contaminated soils were compared, belonging to a wide array of trophic and response levels and using ecoscores for evaluating ecotoxicological and genotoxicological endpoints. The method was applied to four coke factory soils contaminated mainly with PAHs, but also to a lesser extent by heavy metals and cyanides. Aquatic bioassays do not differ from terrestrial bioassays when scaling soils according to toxicity but they are complementary from the viewpoint of ecological relevance. Both aquatic and terrestrial endpoints are strongly correlated with concentrations of 3-ring PAHs. This evaluation procedure allows us to propose a cost-effective battery which embraces a wide array of test organisms and response levels: it includes two rapid bioassays (Microtox(®) and springtail avoidance), a micronucleus test and three bioassays of a longer duration (algal growth, lettuce germination and springtail reproduction). This battery can be recommended for a cost-effective assessment of polluted/remediated soils.

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“…The in vitro micronucleus assay was performed according the procedure described by Nesslany and Marzin (1999). This micro-method used mouse lymphoma cells L5178Y.…”

Section: Aquatic (Geno)toxicity Testsmentioning

confidence: 99%

Comparison of solid and liquid-phase bioassays using ecoscores to assess contaminated soils

Lors

Ponge²,

Aldaya³

et al. 2011

Environmental Pollution

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“…This test consisted in an in vitro micronucleus test using mice lymphoma cells (Nesslany and Marzin, 1999). This test was carried out by Pasteur Institute (Lille, FRANCE).…”

Section: Toxicity Of Soils and Aliquotsmentioning

confidence: 99%

In-Situ Phytoremediation of Pahs Contaminated Soils Following a Bioremediation Treatment

Denys

Rollin

Guillot

et al. 2006

Water Air Soil Pollut: Focus

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Phytoremediation of pollutants in soils is an emerging technology, using different soilplant interaction properties. For organic pollutants, such as polycyclic aromatic hydrocarbons (PAHs), phytodegradation seems to be the most promising approach. It occurs mostly through an increase of the microbial activity in the plant rhizosphere, allowing the degradation of organic substances, a source of carbon for soil microbes. Despite a large amount of available data in the literature concerning laboratory and short term PAH phytodegradation experiments, no actual field application of such technique was previously carried out.In the present study, a soil from a former coking plant was used to evaluate the feasibility and the efficiency of PAH phytodegradation in the field during a three years trial and following a bioremediation treatment. Before the phytoremediation treatment, the soil was homogenized and split into six independent plots with no hydrological connections. On four of these plots, different types of common plant species were sowed: mixture of herbaceous species, short cut (P1), long cut (P2), ornamental plants (P3) and trees (P4). Natural vegetation was allowed to grow on the fifth plot (P5), and the last plot was weeded (P6). Each year, representative sampling of two soil horizons (0-50 and 50-100 cm) was carried out in each plot to characterize the evolution of PAHs concentration in soils and in soils solution obtained by lixiviation. Possible impact of the phytoremediation technique on ecosystems was evaluated using different eco-and genotoxicity tests both on the soil solid matrix and on the soil solution.For each soil horizon, comparable decrease of soil total PAHs concentrations were obtained for three plots, reaching a maximum value of 26% of the initial PAHs concentration. The decrease mostly concerned the 3 rings PAHs. The overall low decrease in PAHs content was linked to a drastic decrease in PAHs availability likely due to the bioremediation treatment. However, soil solutions concentration showed low values and no signficant toxicity was characterized. The mixture of the herbaceous species seemed to be the most promising plants to be used in such procedure.

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“…These characteristics have led to its widespread use as an efficient and relatively simple method to screen drug candidates and other test articles for clastogenic and aneugenic potentials [Nesslany and Marzin, 1999]. Furthermore, there have been concerted efforts to establish robust protocols so that the in vitro micronucleus test can serve as a source of cytogenetic damage information for regulatory submission purposes in place of in vitro chromosome aberration results von der Hude et al, 2000;Garriott et al, 2002;Phelps et al, 2002;KirschVolders et al, 2003].…”

Section: Introductionmentioning

confidence: 99%

In vitro micronucleus scoring by flow cytometry: Differential staining of micronuclei versus apoptotic and necrotic chromatin enhances assay reliability

Avlasevich

Bryce

Cairns

et al. 2005

Environ and Mol Mutagen

140

109

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The in vitro micronucleus test has received considerable attention in recent years for its use in drug safety assessment and toxicological research. The less tedious nature of the assay relative to chromosome aberration analyses is a driving force, and explains why many chemical and drug safety programs have adopted the endpoint. Development of a high-throughput micronucleus scoring system would further enhance the utility of the assay for lead optimization and other early drug development work. Although several variations of a flow cytometric (FCM) method for scoring cell-culture-derived micronuclei (MN) have been described in the literature, they have been unable to distinguish true MN from apoptotic and necrotic chromatin (Nüsse M and Marx K 1997: Mutat Res 392: 109-115). Here, we report advances to this methodology whereby a sequential staining procedure is used to differentially label these types of sub-2n particles. With the use of ethidium monoazide (EMA), the chromatin of dead and dying cells is labeled. After a photoactivation step that covalently binds EMA to chromatin, cytoplasmic membranes are digested and resulting lysates are incubated with RNase plus a pan-nucleic acid dye (SYTOX Green). This process provides a suspension of free nuclei and sub-2n particles that are labeled with either SYTOX or SYTOX and EMA. Preliminary studies with heat-shocked L5178Y mouse cells demonstrated that EMA stains necrotic and mid- through late-stage apoptotic cells. Importantly, the sequential labeling procedure provided reliable micronucleus enumeration, even when cultures contained high percentages of dead cells. Subsequently, experiments with the following diverse genotoxicants were performed: hydroxyurea, methyl methanesulfonate, benzo[a]pyrene, etoposide, cyclophosphamide, and vinblastine. Additionally, the nongenotoxicants sucrose, tributyltin methoxide, and dexamethasone were tested up to 5 mg/ml, or to cytotoxic concentrations. FCM data were found to correspond closely with microscopy-based measurements. Collectively, these data suggest that this sequential EMA/SYTOX staining procedure provides reliable, high-throughput enumeration of in vitro MN.

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A micromethod for the in vitro micronucleus assay

Cited by 54 publications

References 36 publications

Comparison of solid and liquid-phase bioassays using ecoscores to assess contaminated soils

Comparison of solid and liquid-phase bioassays using ecoscores to assess contaminated soils

In-Situ Phytoremediation of Pahs Contaminated Soils Following a Bioremediation Treatment

In vitro micronucleus scoring by flow cytometry: Differential staining of micronuclei versus apoptotic and necrotic chromatin enhances assay reliability

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