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Aplicação do ensaio cometa a estudo de danos ao DNA de robalos, Centropomus parallelus (Poey, 1860), expostos à ß-naftoflavona; Comet Assay applied to DNA damage study in fat snook, Centropomus parallelus (Poey, 1860), exposed to β-naphthoflavone

Di Paolo, Carolina
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Dissertação de Mestrado Formato: application/pdf
Publicado em 01/09/2006 PT
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66.6%
O Ensaio Cometa (Eletroforese em Gel de Célula Única) foi aplicado ao estudo do potencial genotóxico da β-naftoflavona (BNF) em exposição in vivo a eritrócitos de robalos, Centropomus parallelus. Condições específicas para o ensaio cometa de células de sangue de robalos foram estabelecidas com base em informações obtidas em literatura; e através de experimentos com exposição in vitro a diferentes concentrações de peróxido de hidrogênio, e com desenrolamento e eletroforese em diferentes alcalinidades e voltagens. Para avaliar o potencial genotóxico da BNF, os peixes foram expostos a 1ppm e 5ppm de BNF por 24, 48 e 72 horas. Controles foram mantidos em água do mar e em água do mar com DMSO, utilizado com solvente. Células de sangue foram coletadas, submetidas a ensaio cometa em versão alcalina de pH>13 e coradas por prata. Os cometas foram analisados por métodos visuais, incluindo Índice de Danos, Porcentagem de Danos e Freqüência de Danos, e através do sistema de análise de imagem ScionImage. Exposições in vitro de eritrócitos a H2O2 resultaram em relação dose-resposta em pH 12,6 e pH>13, indicando aplicabilidade do ensaio a células de sangue de robalos. Exposições in vivo a BNF indicaram tendência a maior Índice de Danos em grupos expostos comparados a controles...

Teste do Cometa: aplicação ao estudo de vida útil de filés de Tilápia (Oreochromis niloticus - LINNAEUS, 1758); Comet Assay: application in shelf-life study of Tilapia (Oreochromis niloticus - LINNAEUS, 1758) fillets

Pacola, Gian Stefani
Fonte: Biblioteca Digitais de Teses e Dissertações da USP Publicador: Biblioteca Digitais de Teses e Dissertações da USP
Tipo: Dissertação de Mestrado Formato: application/pdf
Publicado em 13/03/2013 PT
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66.66%
O presente estudo teve por objetivo avaliar o desempenho do Teste do Cometa em confronto com métodos convencionais de avaliação de frescor e qualidade higiênica e sanitária de filés de Tilápia (Oreochromis niloticus - LINNAEUS, 1758) refrigerados, durante a vida útil comercial, correlacionando os seus resultados com aqueles obtidos nas análises físico-químicas, microbiológicas e sensorial. As análises foram realizadas nos dias zero - cinco - nove - 12 - 14 pós-processamento dos filés. Em cada dia de estudo foram processados três filés diferentes, pertencentes ao mesmo lote de produto. O protocolo foi repetido em três momentos diferentes, completando 45 amostras. Nos resultados encontrou-se no dia zero e no quinto dia útil do produto, condições impróprias para consumo com relação aos microrganismos psicrotróficos e mesófilos, respectivamente, e o tipo de cometa predominante foi o tipo 3. Aos nove dias de vida útil dos filés, a média dos parâmetros Trimetilamina, Odor e Intenção de Compra indicavam produto impróprio e no teste do cometa predomina o tipo 5. Aos 12 dias de vida útil dos filés de Tilápia, refrigerados, bases voláteis nitrogenadas totais e textura obtiveram resultados impróprios, enquanto que a aparência se revelou imprópria a partir do 14º dia. Concluiu-se que o teste do cometa se presta para o estudo de vida útil de filés de Tilápia refrigerados e apresentou correlação com os métodos convencionais de avaliação de frescor e qualidade higiênica e sanitária do pescado. A partir do 9º dia de estudo predominavam cometas do tipo 5...

Lack of genotoxicity induced by endogenous and synthetic female sex hormones in peripheral blood cells detected by alkaline Comet assay

Braz, Mariana Gobbo; Favero Salvadori, Daisy Maria
Fonte: Wiley-Blackwell Publicador: Wiley-Blackwell
Tipo: Artigo de Revista Científica Formato: 414-420
ENG
Relevância na Pesquisa
66.63%
The etiology of hormone-induced cancers has been considered to be a combination of genotoxic and epigenetic events. Currently, the Comet assay is widely used for detecting genotoxicity because it is relatively simple, sensitive, and capable of detecting various kinds of DNA damage. The present study evaluates the genotoxic potential of endogenous and synthetic sex hormones, as detected by the Comet assay. Blood cells were obtained from 12 nonsmoking and 12 smoking women with regular menstrual cycles and from 12 nonsmoking women taking low-dose oral contraceptives (OC). Peripheral blood samples were collected at three phases of the menstrual cycle (early follicular, mean follicular, and luteal phases), or at three different moments of oral contraceptive intake. Three blood samples were also collected from 12 healthy nonsmoking men, at the same time as oral contraceptive users. Results showed no significant difference in the level of DNA damage among the three moments of the menstrual cycle either in nonsmoking and smoking women, or between them. No significant difference in DNA damage was also observed among oral contraceptive users, nonusers, and men. Together, these data indicate lack of genotoxicity induced by the physiological level of the female sex hormones and OC as assessed by the alkaline Comet assay. In conclusion...

Genotoxicity and mutagenicity of water contaminated with tannery effluents, as evaluated by the micronucleus test and comet assay using the fish Oreochromis niloticus and chromosome aberrations in onion root-tips

Matsumoto, Silvia Tamie; Mantovani, Mário Sérgio; Malaguttii, Mirtis Irene Ariza; Dias, Ana Lúcia; Fonseca, Inês Cristina; Marin-Morales, Maria Aparecida
Fonte: Sociedade Brasileira de Genética Publicador: Sociedade Brasileira de Genética
Tipo: Artigo de Revista Científica Formato: 148-158
ENG
Relevância na Pesquisa
66.53%
Cytotoxicity of metals is important because some metals are potential mutagens able to induce tumors in humans and experimental animals. Chromium can damage DNA in several ways, including DNA double strand breaks (DSBs) which generate chromosomal aberrations, micronucleus formation, sister chromatid exchange, formation of DNA adducts and alterations in DNA replication and transcription. In our study, water samples from three sites in the Córrego dos Bagres stream in the Franca municipality of the Brazilian state of São Paulo were subjected to the comet assay and micronucleus test using erythrocytes from the fish Oreochromis niloticus. Nuclear abnormalities of the erythrocytes included blebbed, notched and lobed nuclei, probably due to genotoxic chromium compounds. The greatest comet assay damage occurred with water from a chromium-containing tannery effluent discharge site, supporting the hypothesis that chromium residues can be genotoxic. The mutagenicity of the water samples was assessed using the onion root-tip cell assay, the most frequent chromosomal abnormalities observed being: c-metaphases, stick chromosome, chromosome breaks and losses, bridged anaphases, multipolar anaphases, and micronucleated and binucleated cells. Onion root-tip cell mutagenicity was highest for water samples containing the highest levels of chromium.

Mutagenic and genotoxic effects of the Atrazine herbicide in Oreochromis niloticus (Perciformes, Cichlidae) detected by the micronuclei test and the comet assay

de Campos, Bruna; de Angelis, Ventura Dejanira de Fransceschi; Marin-Morales, Maria Aparecida
Fonte: Academic Press Inc. Elsevier B.V. Publicador: Academic Press Inc. Elsevier B.V.
Tipo: Artigo de Revista Científica Formato: 42-51
ENG
Relevância na Pesquisa
66.58%
Atrazine is the triazinic herbicide most found in the rural aquatic environments due to its extensive use and its stability in such places. The mutagenicity and the genotoxicity of different concentrations of the Atrazine herbicide were determinated by the micronucleus test and the comet assay, using Oreochromis niloticus as test-system. The tested concentrations of Atrazine herbicide were 6.25, 12.5 and 25 mu g/L, both for the micronuclei test and for the comet assay. The results showed a significant rate of micronuclei and nuclear abnormalities for all the tested concentrations of Atrazine herbicide. For the comet assay, we also observed results significantly different from the control in 6.25, 12.5 and 25 mu g/L concentrations. Due to these results, we could infer that such herbicide may be dangerous to the lives of those organisms exposed to it. (c) 2007 Elsevier B.V. All rights reserved.

Anesthesia of fish with benzocaine does not interfere with comet assay results

De Miranda Cabral Gontijo, Álisson Marques; Barreto, Rodrigo Egydio; Speit, Günter; Valenzuela Reyes, Victor Alexis; Volpato, Gilson Luiz; Favero Salvadori, Daisy Maria
Fonte: Universidade Estadual Paulista Publicador: Universidade Estadual Paulista
Tipo: Artigo de Revista Científica Formato: 165-172
ENG
Relevância na Pesquisa
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Fish blood erythrocytes are frequently used as sentinels in biomonitoring studies. Usually, fish blood is collected by painful cardiac or caudal vein punctures. Previous anesthesia could decrease animal suffering but it is not known at present whether anesthesia can cause confounding effects. Therefore, using the alkaline single cell gel (SCG)/comet assay with blood erythrocytes of the cichlid fish Nile tilapia, we tested for a possible modulation of induced DNA damage (methyl methanesulfonate; MMS) by the anesthetic benzocaine administered by bath exposure (80mg/l for ∼10min). Furthermore, benzocaine (80-600mg/l) was tested for its genotoxic potential on fish erythrocytes in vitro and for potential interactions with two known genotoxins (MMS and hydrogen peroxide). Our results did neither indicate a significant increase in the amount of DNA damage (even after a 48h follow-up), nor indicated interactions with MMS-induced DNA damage when fish were exposed to benzocaine in vivo. There was also no increase in DNA damage after in vitro exposure of fish erythrocytes to benzocaine. Clear concentration-related effects were observed for the two genotoxins in vitro, which were not significantly altered by the presence of benzocaine. These results suggest that anesthesia of fish does not confound comet assay results and the use of blood samples from anesthetized fish can be recommended with regard to animal welfare. © 2002 Elsevier Science B.V. All rights reserved.

Investigation of the Genotoxic Potential of the Waters of a River Receiving Tannery Effluents by Means of the in vitro Comet Assay

Matsumoto, Silvia Tamie; Mantovani, Mário Sérgio; Mallaguti, Mirtis I.; Marin-Morales, Maria Aparecida
Fonte: Universidade Estadual Paulista Publicador: Universidade Estadual Paulista
Tipo: Artigo de Revista Científica Formato: 395-401
ENG
Relevância na Pesquisa
66.55%
The comet assay has been described as an efficient tool for the detection of changes in the DNA molecule of cells exposed to contaminating agents in vivo and in vitro. The possible environmental contamination due to the persistence of chromium residues from tannery effluents was determined in the waters of the Córrego dos Bagres stream, Municipal district of Franca/SP, by the comet assay on CHO-K1 cells. Water samples were collected during the four seasons of the year 2001 at three distinct stations along the river. The data suggest that the comet test showed good sensitivity for the environmental monitoring of these waters and indicated that this test can be efficient for the determination of the quality of waters contaminated with effluents containing heavy metal residues such as chromium.

Lack of genotoxicity of formocresol, paramonochlorophenol, and calcium hydroxide on mammalian cells by comet assay

Ribeiro, Daniel Araki; Marques, Mariãngela Esther Alencar; Salvadori, Daisy Maria Favero
Fonte: Universidade Estadual Paulista Publicador: Universidade Estadual Paulista
Tipo: Artigo de Revista Científica Formato: 593-596
ENG
Relevância na Pesquisa
66.58%
Formocresol, paramonochlorophenol, and calcium hydroxide are widely used in dentistry because of their antibacterial activities in root canal disinfection. However, the results of genotoxicity studies using these materials are inconsistent in literature. The goal of this study was to examine the genotoxic potential of formocresol, paramonochlorophenol, and calcium hydroxide using mouse lymphoma cells and human fibroblasts cells in vitro by the comet assay. Data were assessed by Kruskal-Wallis nonparametric test. The results showed that all compounds tested did not cause DNA damage for the tail moment or tail intensity parameters. These findings suggest that formocresol, paramonochlorophenol, and calcium hydroxide do not promote DNA damage in mammalian cells and that the comet assay is a suitable tool to investigate genotoxicity.

Evaluation of the genotoxic potential due to the action of an effluent contaminated with chromium, by the comet assay in CHO-K1 cultures

Matsumoto, Silvia Tamie; Rigonato, Janaína; Mantovani, Mário Sérgio; Marin-Morales, Maria Aparecida
Fonte: Universidade Estadual Paulista Publicador: Universidade Estadual Paulista
Tipo: Artigo de Revista Científica Formato: 40-46
ENG
Relevância na Pesquisa
66.55%
The comet assay technique has been considered to be more efficient in the biomonitoring of aquatic environments that the micronucleus and sister chromatid exchange techniques. The comet assay has been used to determine breaks in the DNA strands of organisms exposed to pollutants with a genotoxic potential. The comet technique was applied to CHO-K1 cells in order to evaluate the genotoxic potential of the waters of the Sapucaizinho River (Municipality of Patrocínio Paulista, State of São Paulo, Brazil), which receive tannery effluents and therefore are contaminated with chromium. The results indicated high genotoxicity of the waters collected at sites located downstream from the emission of tannery effluents, where the concentration of chromium was found to be high.

Lack of DNA damage induced by fluoride on mouse lymphoma and human fibroblast cells by single cell gel (comet) assay

Ribeiro, Daniel Araki; Alves de Lima, Patrícia Lepage; Marques, Mariângela Esther Alencar; Salvadori, Daisy Maria Favero
Fonte: Universidade Estadual Paulista Publicador: Universidade Estadual Paulista
Tipo: Artigo de Revista Científica Formato: 91-94
ENG
Relevância na Pesquisa
66.58%
Fluoride has widely been used in Dentistry because it is a specific and effective caries prophylactic agent. However, excess fluoride may represent a hazard to human health, especially by causing injury on genetic apparatus. Genotoxicity tests constitute an important part of cancer research for risk assessment of potential carcinogens. In this study, the potential DNA damage associated with exposure to fluoride was assessed by the single cell gel (comet) assay in vitro. Mouse lymphoma and human fibroblast cells were exposed to sodium fluoride (NaF) at final concentration ranging from 7 to 100 μg/mL for 3 h at 37μC. The results pointed out that NaF in all tested concentrations did not contribute to DNA damage as depicted by the mean tail moment and tail intensity for both cellular types assessed. These findings are clinically important because they represent a valuable contribution for evaluation of the potential health risk associated with exposure to agents usually used in dental practice.

Paracoccidioidomycosis: No genetic damage in human peripheral blood cells of patients assessed by single-cell gel (comet) assay

Ribeiro-Vieira, Renata Aparecida Martinez Antunes; Ribeiro, Daniel Araki; Salvadori, Daisy Maria Favero; Marques, Sílvio Alencar
Fonte: Universidade Estadual Paulista Publicador: Universidade Estadual Paulista
Tipo: Artigo de Revista Científica Formato: 476-478
ENG
Relevância na Pesquisa
66.58%
Paracoccidioidomycosis is a systemic fungal infection caused by Paracoccidioides brasiliensis. As infectious diseases can cause DNA damage, the authors aimed at analyzing DNA breakage in peripheral blood cells of patients with paracoccidioidomycosis by using the comet assay. The results suggested that paracoccidioidomycosis does not cause genotoxicity.

Influence of endogenous and synthetic female sex hormones on human blood cells in vitro studied with comet assay

Braz, Mariana Gobbo; Fávero Salvadori, Daisy Maria
Fonte: Universidade Estadual Paulista Publicador: Universidade Estadual Paulista
Tipo: Artigo de Revista Científica Formato: 972-976
ENG
Relevância na Pesquisa
66.67%
The comet assay has been conducted with numerous cell lines to assess in vitro genotoxicity. In order to use the comet assay as part of an in vitro test for evaluating genotoxicity, however, there are cell-specific factors that need to be better understood. In this present study we have evaluated some factors that may impact upon the DNA damage detected in whole blood (WB) cells and lymphocytes (ILs). Experiments were conducted comparing responses of both cells, and investigating the effects of the female hormonal cycle, and oral contraceptive (OC) use on DNA damage detection in the in vitro comet assay, at three sampling time. No significant differences were detected in the basal levels of DNA damage detected in ILs and WB cells from women OC users and non-users and from men. Basal DNA damage in ILs was unaffected by gender and stage of the menstrual cycle or the stage of the treatment schedule. Our results also indicated that the H2O2 induces DNA damage in human lymphocytes independently of gender, low-dose OC use and hormonal fluctuation. However, data showed that in 3rd sampling of menstrual cycle, lymphocytes were more resistant to H2O2-induced DNA damage than those from OC users and men. © 2007 Elsevier Ltd. All rights reserved.

Mutagenic effects of tributyltin and inorganic lead (Pb II) on the fish H. malabaricus as evaluated using the comet assay and the piscine micronucleus and chromosome aberration tests

Ferraro,Marcos Vinícius M.; Fenocchio,Alberto Sérgio; Mantovani,Mario Sérgio; Ribeiro,Ciro de Oliveira; Cestari,Marta Margarete
Fonte: Sociedade Brasileira de Genética Publicador: Sociedade Brasileira de Genética
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/01/2004 EN
Relevância na Pesquisa
66.6%
Genotoxicity studies on toxic metals and their organic compounds are very important, especially so in the investigation of the effects of these compounds on the aquatic environments where they tend to accumulate. The use of endemic aquatic organisms as biological sentinels has proved useful to environmental monitoring. We assessed the mutagenic potential of tributyltin (TBT) and inorganic lead (PbII) using samples of the fish Hoplias malabaricus (commonly called traíra) using the comet assay and the piscine micronucleus and chromosome aberration tests. Eighteen H. malabaricus were acclimatized in three individual aquariums, each containing six fish, six fish being exposed to 0.3 mg/g of body weight (bw) of TBT, six to 21 mg/g bw of PbII and six being used as controls. Exposure to TBT and PbII was achieved by feeding the fish every five days with Astyanax (a small fish that is part of the normal diet of H. malabaricus) which had been injected with solutions of TBT, PbII or with water (the control group). After two months the H. malabaricus were sacrificed and their peripheral blood collected and subjected to the comet and micronucleus assays, the chromosome aberration assay being conducted using kidney-tissue. Although the comet assay showed now mutagenic effects at the lead concentrations used but encountered results with TBT...

First genotoxicity study of Paraná river water from Argentina using cells from the clam Corbicula fluminea (Veneroida Corbiculidae) and Chinese hamster (Cricetulus griseus Rodentia, Cricetidae) K1 cells in the comet assay

Caffetti,Jacqueline D.; Mantovani,Mário S.; Pastori,María C.; Fenocchio,Alberto S.
Fonte: Sociedade Brasileira de Genética Publicador: Sociedade Brasileira de Genética
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/01/2008 EN
Relevância na Pesquisa
66.58%
High concentrations of xenobiotics from urban and industrial wastes have contributed to the contamination of many aquatic environments. We used the comet assay to evaluate the genotoxic potential of water collected from the River Paraná, which receives a great deal of waste, at three points (Puerto Piray, Eldorado and Montecarlo) in the Misiones Province of Argentina. The in vivo comet assay used 40 freshwater clams (Corbicula fluminea) while the in vitro comet assay used Chinese hamster (Cricetulus griseus) K1 cell (CHO-K1) cultures with the mutagen ethyl methanesulfonate (EMS) as the positive control and phosphate buffered saline (PBS) as the negative control. Both assays showed statistically significant differences between the three sampling sites in relation to the negative control, the results of this preliminary study indicating that at these three sites water from the Paraná River presents genotoxic potential.

Estimates of DNA damage by the comet assay in the direct-developing frog Eleutherodactylus johnstonei (Anura, Eleutherodactylidae)

Valencia,Laura Carolina; García,Adriana; Ramírez-Pinilla,Martha Patricia; Fuentes,Jorge Luis
Fonte: Sociedade Brasileira de Genética Publicador: Sociedade Brasileira de Genética
Tipo: Artigo de Revista Científica Formato: text/html
Publicado em 01/01/2011 EN
Relevância na Pesquisa
66.63%
The aim of this study was to use the Comet assay to assess genetic damage in the direct-developing frog Eleutherodactylus johnstonei. A DNA diffusion assay was used to evaluate the effectiveness of alkaline, enzymatic and alkaline/enzymatic treatments for lysing E. johnstonei blood cells and to determine the amount of DNA strand breakage associated with apoptosis and necrosis. Cell sensitivity to the mutagens bleomycin (BLM) and 4-nitroquinoline-1-oxide (4NQO) was also assessed using the Comet assay, as was the assay reproducibility. Alkaline treatment did not lyse the cytoplasmic and nuclear membranes of E. johnstonei blood cells, whereas enzymatic digestion with proteinase K (40 !g/mL) yielded naked nuclei. The contribution of apoptosis and necrosis (assessed by the DNA diffusion assay) to DNA damage was estimated to range from 0% to 8%. BLM and 4NQO induced DNA damage in E. johnstonei blood cells at different concentrations and exposure times. Dose-effect curves with both mutagens were highly reproducible and showed consistently low coefficients of variation (CV < 10%). The results are discussed with regard to the potential use of the modified Comet assay for assessing the exposure of E. johnstonei to herbicides in ecotoxicological studies.

The relationship between environmental exposures to phthalates and DNA damage in human sperm using the neutral comet assay.

Singh, Narendra P; Silva, Manori J; Barr, Dana B; Brock, John W; Duty, Susan M.; Ryan, Louise Marie; Herrick, Robert F.; Christiani, David C.; Hauser, Russ B.
Fonte: Harvard University Publicador: Harvard University
Tipo: Artigo de Revista Científica
EN_US
Relevância na Pesquisa
66.66%
Phthalates are industrial chemicals widely used in many commercial applications. The general population is exposed to phthalates through consumer products as well as through diet and medical treatments. To determine whether environmental levels of phthalates are associated with altered DNA integrity in human sperm, we selected a population without identified sources of exposure to phthalates. One hundred sixty-eight subjects recruited from the Massachusetts General Hospital Andrology Laboratory provided a semen and a urine sample. Eight phthalate metabolites were measured in urine by using high-performance liquid chromatography and tandem mass spectrometry; data were corrected for urine dilution by adjusting for specific gravity. The neutral single-cell microgel electrophoresis assay (comet assay) was used to measure DNA integrity in sperm. VisComet image analysis software was used to measure comet extent, a measure of total comet length (micrometers); percent DNA in tail (tail%), a measure of the proportion of total DNA present in the comet tail; and tail distributed moment (TDM), an integrated measure of length and intensity (micrometers). For an interquartile range increase in specific gravity-adjusted monoethyl phthalate (MEP) level...

Application of public-domain statistical analysis software for evaluation and comparison of comet assay data

Verde, Pablo Emílio; Monserrat, Laura Alicia Geracitano; Amado, Lílian Lund; Rosa, Carlos Eduardo da; Bianchini, Adalto; Monserrat, José María
Fonte: Universidade Federal do Rio Grande Publicador: Universidade Federal do Rio Grande
Tipo: Artigo de Revista Científica
ENG
Relevância na Pesquisa
66.69%
A novel approach for statistical analysis of comet assay data (i.e.: tail moment) is proposed, employing public-domain statistical software, the R system. The analytical strategy takes into account that the distribution of comet assay data, like the tail moment, is usually skewed and do not follow a normal distribution. Probability distributions used to model comet assay data included: the Weibull, the exponential, the logistic, the normal, the log normal and log-logistic distribution. In this approach it was also considered that heterogeneity observed among experimental units is a random feature of the comet assay data. This statistical model can be characterized with a location parameter mij, a scale parameter r and a between experimental units variability parameter θ. In the logarithmic scale, the parameter mij depends additively on treatment and random effects, as follows: log(mij) = a0 + a1xij + bi, where exp(a0) represents approximately the mean value of the control group, exp(a1) can be interpreted as the relative risk of damage with respect to the control group, xij is an indicator of experimental group and exp(bi) is the individual risk effects assume to follows a Gamma distribution with mean 1 and variance θ. Model selection is based on Akaike’s information criteria (AIC). Real data coming from comet analysis of blood samples taken from the flounder Paralichtys orbignyanus (Teleostei: Paralichtyidae) and from samples of cells suspension obtained from the estuarine polychaeta Laeonereis acuta (Nereididae) were employed. This statistical approach showed that the comet assay data should be analyzed under a modeling framework that take into account the important features of these measurements. Model selection and heterogeneity between experimental units play central points in the analysis of these data.

Evaluation of the genotoxicity of waters impacted by domestic and industrial effluents of a highly industrialized region of So Paulo State, Brazil, by the comet assay in HTC cells

Manzano, Barbara Cassu; Roberto, Matheus Mantuanelli; Hoshina, Marcia Miyuki; Menegario, Amauri Antonio; Marin-Morales, Maria Aparecida
Fonte: Springer Publicador: Springer
Tipo: Artigo de Revista Científica Formato: 1399-1407
ENG
Relevância na Pesquisa
66.58%
Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES); The problems that most affect the quality of the waters of rivers and lakes are associated with the discharges performed in these environments, mainly industrial and domestic effluents inappropriately treated or untreated. The comet assay is a sensitive tool and is recommended for studies of environmental biomonitoring, which aim to determine the genotoxicity potential of water pollutants. This study aimed to assess the genotoxic potential of the Ribeiro Tatu waters, region of Limeira, So Paulo (SP), by the comet assay with mammalian cells (hepatoma tissue culture (HTC)). Water samples were collected along the Ribeiro Tatu at three distinct periods: November 2008, February 2009 and August 2009, and five collection sites were established: P1, source of the stream; P2, site located downstream the urban perimeter of the municipality of Cordeirpolis and after receiving the pollution load of this city; P3, collection site located upstream the urban perimeter of the city of Limeira; P4, urban area of Limeira; and P5, rural area of Limeira, downstream the discharges of the city sewage. The results showed that for the November 2008 collection, there was no water sample-induced genotoxicity; for the February 2009 collection...

Novel method for the high-throughput processing of slides for the comet assay

Karbaschi, Mahsa; Cooke, Marcus
Fonte: FIU Digital Commons Publicador: FIU Digital Commons
Tipo: Artigo de Revista Científica Formato: application/pdf
Relevância na Pesquisa
66.68%
Single cell gel electrophoresis (the comet assay), continues to gain popularity as a means of assessing DNA damage. However, the assay’s low sample throughput and laborious sample workup procedure are limiting factors to its application. “Scoring”, or individually determining DNA damage levels in 50 cells per treatment, is time-consuming, but with the advent of high-throughput scoring, the limitation is now the ability to process significant numbers of comet slides. We have developed a novel method by which multiple slides may be manipulated, and undergo electrophoresis, in batches of 25 rather than individually and, importantly, retains the use of standard microscope comet slides, which are the assay convention. This decreases assay time by 60%, and benefits from an electrophoresis tank with a substantially smaller footprint, and more uniform orientation of gels during electrophoresis. Our high-throughput variant of the comet assay greatly increases the number of samples analysed, decreases assay time, number of individual slide manipulations, reagent requirements and risk of damage to slides. The compact nature of the electrophoresis tank is of particular benefit to laboratories where bench space is at a premium. This novel approach is a significant advance on the current comet assay procedure.

Studies of DNA repair after an oxidative challenge and its induction by natural compounds : comet assay in Saccharomyces cerevisiae

Azevedo, F.; Johansson, Björn; Oliveira, Rui Pedro Soares de
Fonte: Universidade do Minho Publicador: Universidade do Minho
Tipo: Conferência ou Objeto de Conferência
Publicado em //2007 ENG
Relevância na Pesquisa
66.66%
A reliable and sensitive technique for detection of DNA damage is the Single Cell Gel Assay (also known as comet assay). In this procedure, single cells are exposed to an electric field and damaged DNA moves out of the nucleus, which can be visualized by ethidium bromide staining. Cells display a comet appearance under fluorescence microscopy, the size of the DNA “tail” of the comet being in direct correlation with the extension of DNA damage. In this work we have optimized the comet assay using Saccharomyces cerevisiae cells in order to assess DNA damage caused by oxidative stress. We show that genomic DNA damage increases as the yeast population enters the post-diauxic phase. In addition, the tail comet length decreases by incubation of yeast cells with natural compounds, known to protect cells against oxidative stress (quercetin, ursolic acid), prior to oxidative challenge. Further experiments are being conducted to investigate the DNA repair kinetics by measuring the decrease in the length of the comet tail in a recovery period after an oxidative challenge. We are using mutants affected in RAD1 (an endonuclease of the nucleotide excision repair pathway) and in APN1 (a nuclease of the base excision repair pathway) to determine the repair pathway involved in the oxidative damage. In conclusion...