• 한국환경성돌연변이발암원학회지
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• 제17권2호
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• pp.71-77
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• 1997

The single cell gel electrophoressis(SCGE) assay, also known as the comet assay, is a rapid, simple, visual and sensitive technique for measuring and analysing DNA breakage in mammalian cells. The SCGE or comet assay is a promising test for the detection of DNA damage and repair in individnal cells. It has widespread potential applications in DNA damage and repair studies, genotoxicity testing and biomonitoring. In this microgel electrophoresis technique, cells are embedded in agarose gel on microscope slides, iysed and electrophoresed under alkaline conditions. Cells with increased DNA damage display increased migration of DNA from the nucleus towards the anode. The length of DNA migration indicates the amount of DNA breakage in the cell. The comet assay is also capable of identifying apoptotic cells which contain highly fragmented DNA. Here we review the development of the SCGE assay, existing protocols for the detection and analysis of comets, the relevant underlying principles determining the behaviour of DNA and the potential applications of the technique.

• Journal of Nutrition and Health
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• 제35권2호
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• pp.213-222
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• 2002

The alkaline comet assay has been used with increasing popularity to investigate the level of DNA damage in biomonitoring studies within the last decade in Western countries. The purpose of this study was to evaluate the usefulness of the alkaline comet assay as a biomarker of oxidative DNA damage for monitoring in the Korean population, and also to evaluate the effect of nutritional status and lifestyle factors on H2O2 induced oxidative DNA damage measured by the alkaline comet assay in human lymphocytes. The study population consisted of 61 healthy Korean male volunteers, aged 20-28. Epidemiological background data including dietary habits, smoking habits and anthropometrical measurements were collected through personal interviews. After blood collection, the comet assay in peripheral lymphocytes and plasma lipids analysis was carried out and the results analyzed. Tail moment (TM) and tail length (TL) of the comet assay were use\ulcorner to measure DNA damage in the lymphocytes of the subjects. Statistically significant (p < 0.05) positive correlations were observed between DNA damage (TM or TL) and smoking habits expressed as cigarettes smoked per day and pack years (r = 0.311 and 0.382 for TM, r = 0.294 and 0.350 for TL, respectively). There were also significant positive correlations between DNA damage parameter and waist-hip ratio. Higher plasma triglyceride levels were associated with increased damage to DNA. There were no correlations between the consumption frequencies of vegetables and DNA damage to the subjects. However, consumption frequencies of fruit and fruit juice intake were inversely associated with the TM and TL. The results indicate that die comet assay is a simple, rapid and sensitive method for detecting lymphocyte DNA damage induced by cigarette smoking. Consumption of fruit or fruit juices could potentiall modify the damaged DNA in the human peripheral lymphocytes of young Korean men.

• 한국식품영양과학회지
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• 제37권4호
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• pp.472-476
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• 2008

국내에서 주로 소비되는 채소류와 곡물류의 방사선조사 여부를 신속하게 검지하기 위하여 DNA comet assay의 활용 가능성을 조사하였다. 식품의 종류에 따라 세포용출시간, 세포현탁액의 정치시간 및 세포용해시간 등의 DNA comet assay 조건을 달리하여야 DNA comet이 관찰되는 것으로 나타났다. 전반적으로 방사선조사선량에 따라 comet assay 값이 증가하였는데 유의적 차이는 식품의 종류에 따라 달리 나타났다. 즉, 파는 2 kGy, 마늘은 3 kGy, 토마토는 1 kGy, 쌀가루는 9 kGy, 그리고 서리태는 3 kGy에서 방사선조사 여부에 대한 검지가 가능한 것으로 나타났다. 식물세포는 동물세포에 비해 외부의 영향에 민감하게 작용하기 때문에 DNA도 쉽게 손상되어 크기뿐만 아니라 핵의 모양도 다양하게 나타나는 것으로 보고된 바와 같이 DNA comet assay를 다양한 식품의 방사선조사 신속검지법으로 활용하기 위해서는 식품의 종류에 따른 표준화된 comet 분석조건 및 통계 분석법에 대한 가이드라인 설정이 필요한 것으로 나타났다.

• 한국환경독성학회:학술대회논문집
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• 한국환경독성학회 1997년도 제20회 화학물질의 환경독성과 건강영향
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• pp.101-101
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• 1997

Comet assay, single cell gel electrophoresis has been known as useful, rapid, simple, visual, and sensitive technique for measuring the DNA breakage in mammalian ce1ls. For evaluation of DNA damage using comet assay, early studies reported a change in comet length and intensity with DNA damage using simple visual technique, such as fluorescence microscopy with eyespiece. In recent, some workers are observing and analyzing nucleotide of comets using quantitative fluorescence image analysis system to estimate 'tail moment', which is defined as the product of the tail length and the fraction of total DNA in tail. Our laboratory also adopted the image analysis software for qualification. In addition, many of the practical features of comet assay render it potentially attractive as useful tool for molecular toxicology and carcinogenesis, because the system is already showing considerable promise as rapid predictor in both in vitro and in vivo experimental designs. Recently, the comet assay becomes a attractive technique to study of apoptosis, because apoptotic fragmentation of nuclear DNA into nucleosomal sizes can be evaluated by the comet assay. So, we attempted to apply the comet assay to studying the effect of various stress on the apoptosis-sensitive cell lines. Particularly, focusing on the hyperthermic apoptosis, we could find that heat shock(44˚C for 60 minutes) was sufficient to induced apoptosis in these cell lines. But using the highly sensitive comet assay, we could not detect DNA breaks immediately after heat shock.

• 한국식품영양과학회지
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• 제29권5호
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• pp.843-848
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• 2000

콩류의 방사선 조사 여부를 DNA comet assay로 확인하였다. 대두, 강남콩, 그리고 팥을 각각 0.1, 0.3, 0.5, 0.7, 그리고 1.0 kGy의 저선량으로 조사하고 비조사 시료와 조사시료간의 DNA 손상정도를 현미경으로 육안 검사 및 comet의 tail 길이를 측정하여 상호 비교하였다. 모든 시료에서 비조사 시료보다 조사시료의 tail 길이가 더 길었으며 조사 선량이 증가할수록 tail 길이가 길게 나타났다. 비조사 시료에서도 손상된 세포의 comet 모양의 핵이 관찰되기는 하였으나 대부분 비손상된 세포의 원형모양의 핵이 관찰되었으며 조사된 시료에서는 comet 모양의 핵이 주로 관찰되는 경향이어서 비조사 시료와 조사 시료간에 comet 양상을 비교할 수 있었다. 한편 100개의 세포를 무작위로 선택하여 comet tail 길이를 측정하고 통계분한 결과, 비조사 시료와 조사시료간의 유의적인 차이가 있었고 조사선량이 증가할수록 tail 길이가 유의적으로 증가함을 확인하였다. DNA comet assay로 간단하고 비교적 저렴한 비용을 짧은 시간 내에 콩류의 방사선 조사 여부를 확인할 수 있었다.

• 한국식품영양과학회지
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• 제29권6호
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• pp.1025-1029
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• 2000

방사선 조사된 쇠고기와 돼지고기의 방사선 조사 여부를 판별하는데 DNA comet assay의 활용 가능성을 검토하였다. 쇠고기와 돼지고기는 Co-60 동위 원소를 조사원으로 하여 0.1, 0.3, 0.5, 0.7, 1.0 kGy의 총흡수선량( $\pm$ 5.0%)이 되도록 조사하여, 냉동상태로 보관하였다. 시료로부터 분리 된 세포는 agarose gel과 혼합하여 슬라이드에 깔아주고, lysis 및 전기영동을 하였다. 방사선 조사된 시료의 경우 세포로부터 끌려나오는 DNA절편들은 양극을 방향으로 tail이 형성되었고, 비조사 시료의 경우 tail이 없거나, 일부에서만 작은 tail이 관찰되었다. 방사선 조사유무는 0.1 kGy부터 현미경상으로 검지가 가능하였고, 선량간 차이의 유의성 여부는 통계분석을 통하여 검지가 가능하여, 쇠고기와 돼지고기의 신속한 검지 방법으로 DNA comet assay를 활용할 수 있을 것이다.

• Toxicological Research
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• 제22권4호
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• pp.423-429
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• 2006

Recent reports on the photocarcinogenicity and photogerotoxicity of many compounds led to an increasing awareness for the need of a standard approach to test for photogenotoxicity. The comet assay has been recently validated as a sensitive and specific test system for the quantification of DNA damage. Thus, the objectives of this study are to investigate the utility of photo-comet assay for detecting photo-mutagens, and to evaluate its ability to predict rodent photo-carcinogenicity. Photo-comet assays were performed using L5178Y $Tk^{+/-}$ mouse lymphoma cells on five test substances (8-methoxypsoralen, chlorpromazine, lomefloxacin, anthracene and retinoic acid) that demonstrated positive results in photocarcinogenicity tests. For the best discrimination between the test substance-mediated DNA damage and the undesirable DNA damage caused by direct UV absorption, a UV dose-response of the cells in the absence of the test substances was firstly fnalized. Out of 5 test substances, positive comet results were obtained for chlorpromazine, lomefloxacin, anthracene and retinoic acid while 8-methoxypsoralen found negative. An investigation into the predictive value of this photo-comet assay for determining the photocarcinogenicity showed that photo-comet assay has relatively high sensitivity. Therefore, the photo-comet assay with mammalian cells seems to be a good and sensitive predictor of the photocarcinogenic potential of new substances.

• 한국임상수의학회지
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• 제13권2호
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• pp.158-162
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• 1996

Comet assay 는 포유류 세포에서 DNA의 파괴를 측정하는데 있어 신속하고 단순하며 시각적이고 민감한 방법이다. Apoptosis에서는 세포핵의 광범위한 DNA의 붕괴가 일어나므로 comet assay는 종양세포에서 apoptosis가 발생되었는가를 알아내는데 유용하다. 본 연구는 apoptosis 연구의 결과가 변화되지 않도록 comet assay slides를 좀 더 오래 보관할 수 있는 방법을 개발하고자 시행되었다. 개의 종양세포를 가지고 alkali comet assay를 끝낸 뒤 slides를 진공 건조기에서 꺼내서 증류수로 점적하여 10-20분간 침수시키고 현광현미경하에서 육안적으로 관찰하였다. 건조후 3-4일, 1주, 2주, 3주, 4주 및 7주의 slides에서 apoptosis 회복율(%)은 각각 98.1, 98.3, 99.4, 80.8 및 35.2%이었다. 3주 이내의 slides에서 대조군과 비교하여 apoptosis 회복율에서 차이가 없었으나 4주 이상의 slides를 건조후 침수시키는 방법을 이용하였을 때 apoptosis 평가에서 건조 후 3주간까지는 처음의 결과와 차이가 없으며, 이 방법을 이용하여 comet slide의 좀 더 긴 기간이 보관과 보관후의 재평가에서 이용될 수 있는 좋은 방법이 된다.

• Ocean and Polar Research
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• 제33권2호
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• pp.127-133
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• 2011

The comet assay, also called single-cell electrophoresis (SCGE) assay, is a potential sensitive monitoring tool for DNA damage in cells. The primary objective of this study was to use comet assay to ascertain if the blood cells of flounder (Pleuronichthys olivaceus) and muscle cells of clam (Saxidomus purpurata) are suitable for genotoxicity screening. This was achieved by initially exposing blood and muscle cells under in vitro conditions to the reference genotoxin hydrogen peroxide ($H_2O_2$); strong correlation between $H_2O_2$ concentration and comet values were found. Subsequently, the identification of DNA damage in isolated cells from flounder and clam was performed under in vivo exposure to benzo(a)pyrene (BaP) and tributyltin (TBT). Flounder and clam were exposed to different concentrations (1, 10, 50, 100 ${\mu}g/L$) of BaP or TBT for 4 days. Regardless of treated chemicals, blood cells of flounder were more prone to DNA breakage compared to muscle cells of clam. In conclusion, in vivo genotoxicity of BaP and TBT can be biomonitored using the comet assay. This study suggests that flounder and clam do show potential as mediums for monitoring genotoxic damage by comet assay.

• 한국수산과학회지
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• 제36권4호
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• pp.346-351
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• 2003

Single-cell gel electrophoresis (comet assay) was used to detect DNA single strand break in blood cells from several marine fish species. Three fish species were collected from Georgia coastal area. Mummichog, Fundulus heteroclitus showed higher DNA damage than sea bass, Lateolabrax japonicus and trout, Oncorhynchus masou masou under the same experimental conditions. Mummichogs had more alkaline-labile sites on their DNA than other fish species. The comet assay with mummichog blood cells at pH 12.5 showed a dose-response curve with the increasing concentrations of hydrogen peroxide. While the isolated leucocytes showed no increase of DNA damage after in vitro exposure to 2-methyl-1,4-naphthoquinone (MNQ), erythrocytes showed dose-dependent DNA damage. These results indicate that the comet assay can be applied successfully as a bioassay using erythrocyte for environmental monitoring.