单细胞凝胶电泳诱变检测系统的研究及应用
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摘要
本研究将细胞培养和单细胞凝胶电泳结合起来建立了单细胞凝胶电泳诱变检测系统，并用该检测系统检测兽药添加剂喹烯酮和喹乙醇对DNA的损伤程度。并基于阳性致突变物H_2O_2作用于非洲绿猴肾Vero细胞引起细胞DNA损伤的原理，研究了其关键步骤-裂解时间，以9.1～273μmol／L剂量的H_2O_2染毒处于对数生长期的Vero细胞3h后，收获细胞用于制备三明治凝胶板，分别裂解1h、2h、3h和4h并选择最适裂解时间。裂解后的细胞进行解旋与电泳、中和、DNA染色和荧光检测，通过分析彗星样细胞发生率和彗尾的长短等指标，证明裂解时间以2～3h为宜。
应用建立的单细胞凝胶电泳诱变检测系统检测喹烯酮和喹乙醇对DNA的损伤程度。喹烯酮染毒剂量在1～5μg／ml时细胞存活率＞85％，以1～5μg／ml剂量的喹烯酮染毒处于对数生长期的Vero细胞，后经过电泳，通过分析彗星样细胞发生率和彗尾长短等指标，结果表明染毒剂量在2μg／ml对DNA造成中度损伤。喹乙醇染毒剂量在2～10μg／ml时细胞存活率＞90％，以2～10μg／ml剂量的喹乙醇染毒处于对数生长期的Vero细胞，后经过电泳，通过分析彗星样细胞发生率和彗尾长短等指标，结果表明染毒剂量在6μg／ml对DNA造成中度损伤。本试验表明喹烯酮和喹乙醇具有致突变作用。
本论文首次将细胞培养技术和单细胞凝胶电泳结合起米应用于检测兽药对DNA的损伤，并对该方法中的一些关键技术条件进行了研究，为兽药安全评价体系的更新提供了一定的技术贮备。
We applied single cell gel electrophoresis and cell culture technique, which constitute sing cell gel electrophoresis assay system for detecting mutagenicity to detect mutagenesis in vitro induced by animal drug Quinocetone and Olaquindox. And confirmed optimum lysing-time. Vero cells in the period of logarithm- growth time were treated with 9.1~273u mol/L H2O2 at 37℃ 3h, then were lysed for lh, 2h, 3h and 4h to find optimum lysing-time. Index -percentage of "comet" cell and "comet
tail" were analysed that indicated the optimum time is 2~3h.
The degree of DNA damage by Quinocetone and Olaquindox were detected by SCGE assay, when Vero cells were treated with 1~5 ug/ml Quinocetone the percentage of live cells was above 85% and with 2-10ug/ml Olaquindox the percentage of live cells was above 90%. Vero cells in the period of logarithm- growth time were treated with 1~5ug/ml Quinocetone or 2~10u g/ml Olaquindox at 37℃ for 3h, and were lysed at 4℃ for 3h. Then were electrophored. The extent of DNA migration were measured. Index -percentage of "comet" cell and "comet tail" were analysed that indicated when Vero cells were treated with 2 u g/ml Quinocetone Vero cells got midium-grade damage, and were treated with 6 u g/ml Olaquindox Vero cells got midium-grade damage. The experimentation indicated that Quinocetone and Olaquindox can cause cellular mutation.
In this paper cell culture and SCGE were applied first to detect DNA damage worked by animal drugs. And pivotal coditions is studyed.This affords technic-reservior for renewal of safety evaluation system of animal drugs.
应用建立的单细胞凝胶电泳诱变检测系统检测喹烯酮和喹乙醇对DNA的损伤程度。喹烯酮染毒剂量在1～5μg／ml时细胞存活率＞85％，以1～5μg／ml剂量的喹烯酮染毒处于对数生长期的Vero细胞，后经过电泳，通过分析彗星样细胞发生率和彗尾长短等指标，结果表明染毒剂量在2μg／ml对DNA造成中度损伤。喹乙醇染毒剂量在2～10μg／ml时细胞存活率＞90％，以2～10μg／ml剂量的喹乙醇染毒处于对数生长期的Vero细胞，后经过电泳，通过分析彗星样细胞发生率和彗尾长短等指标，结果表明染毒剂量在6μg／ml对DNA造成中度损伤。本试验表明喹烯酮和喹乙醇具有致突变作用。
本论文首次将细胞培养技术和单细胞凝胶电泳结合起米应用于检测兽药对DNA的损伤，并对该方法中的一些关键技术条件进行了研究，为兽药安全评价体系的更新提供了一定的技术贮备。
We applied single cell gel electrophoresis and cell culture technique, which constitute sing cell gel electrophoresis assay system for detecting mutagenicity to detect mutagenesis in vitro induced by animal drug Quinocetone and Olaquindox. And confirmed optimum lysing-time. Vero cells in the period of logarithm- growth time were treated with 9.1~273u mol/L H2O2 at 37℃ 3h, then were lysed for lh, 2h, 3h and 4h to find optimum lysing-time. Index -percentage of "comet" cell and "comet
tail" were analysed that indicated the optimum time is 2~3h.
The degree of DNA damage by Quinocetone and Olaquindox were detected by SCGE assay, when Vero cells were treated with 1~5 ug/ml Quinocetone the percentage of live cells was above 85% and with 2-10ug/ml Olaquindox the percentage of live cells was above 90%. Vero cells in the period of logarithm- growth time were treated with 1~5ug/ml Quinocetone or 2~10u g/ml Olaquindox at 37℃ for 3h, and were lysed at 4℃ for 3h. Then were electrophored. The extent of DNA migration were measured. Index -percentage of "comet" cell and "comet tail" were analysed that indicated when Vero cells were treated with 2 u g/ml Quinocetone Vero cells got midium-grade damage, and were treated with 6 u g/ml Olaquindox Vero cells got midium-grade damage. The experimentation indicated that Quinocetone and Olaquindox can cause cellular mutation.
In this paper cell culture and SCGE were applied first to detect DNA damage worked by animal drugs. And pivotal coditions is studyed.This affords technic-reservior for renewal of safety evaluation system of animal drugs.
引文
1. Ames, et al, Dietary carcinogens and anticarcinogens, Science, 1983, 221:1256-1264.
2. Betti C,Banath TP, Durand RE, Comparative studies by Comet test and SCGE analysis in human lymphocytes from 200 healthy subjects, [J]. Mutation Research,1994,306:323-333.
3. Betti C, DaviniT, GiannessiL, et al, Comparative studies by Comet test and SCGE analysis in human lymphocytes from 200 healthy subjects, [J]. Mutation Research, 1995,343:201-207.
4. Betti C, et al, Microgel electrophoresis assay (comet test) and SCG analysis in human lymphocytes from 100 normal subjects. Mutat. Res., 1994,307:323-333.
5. Cerutli P.A., Prooxidant states and tumor promotion. Science,1985,227:375-381.
6. Anderson D.et al, The effect of various antioxidants and other modifying agents on oxygen-radical-generated DNA damage in human lymphocytes in the COMET assay, Mutation Research, 1994,307:261-271
7. Daryl W. Fairbairn et al,The cometassay:a comprehensive review, Mutation Research,1995,339:37-59
8. De Meo, M. Laget et al, Genotoxic activity of potassium permangannate in acidic solution,Mutation Research,1991,260:295-306
9. Duthie SJ, et al, The effect of dietary flavonoida on DNA damage (strand breaks and oxidized pyrimdines) and growth in human cells. Mutat. Res.,1997,390:141-151.
10. Gedik CM, et ai, Single cell electrophoresis applied to the analysis of UV-C damage and its repair in human cells, Int. J Radiat. Biol, 1992,62:313
11. Gellik, C.M., S.W.B. Ewen and A.R. Collins, Single-cell gel electrophoresis applied to the analysis of UV-C damage and its repair in human cells, Int.J Radiat. Biol.,1992, 62:313-320
12. Green, M.H.L. et ai, UV-C sensitivity of unstimulated and stimulated human lymphocytes from normal and xeroderma pigmentosum donors in the comet assay: A potential diagnostic technique,Mutation Research, 1992, 273:137-144
13. Grichuer T, et al, Induction of somatic DNA damage as measure by single cell gel electrophoresis and point mutation in leaves of tobacco plants. Mutat. Res., 1998,401 (2):143-152
14. Gutierrez S,et al,Application of the single tee gel electrophoresis(SCGE) assaytothe detected of DNA damage induced by 1311 treatment in hyperthyroidism patients. Mutagenesis, 1998,13:95 98.
15. Guyton, et al, Oxidative mechanisms in carcinogenesis, Br. Med. Bull., 1993, 49:523-544
16. Heliman B, et al, the conpects of tail moment and tail inertia in the single cell gel electrophoresis assay. Murat. Res., 1995,336:123-131
17. Lubin JH, Blot WJ, Berrino FM, odifying risk of developing lung cancer by changing habits of cigarette smoking, [J].BrMedJ, 1984,288:1953-1956
18. Maarit HL, et al, Repair of γ-irradiation-induced DNA single-strand breaks in human
bonemarrow cells: Effects of a second irradiation. Mutat. Res., 1997, 388:33-44.
19. Maes A, et al, Cytogenetic effects of 935-2 MH_2 (GSM) microwaves a lone and in combition with mitomycinC, Mutat. Res., 1997,393(1-2):151
20. Malyapa R A, et al, DNA damage in rat brain cells after in vivo exposure to 2450 MH2 electromagnetic radiation and various methods of euthanasia. Radiat Res.,1998,149(6):637-645
21..Green M.H.L et al, UV-C sensitivity of unstimulated and stimulated human lymphocytes from normal and xeroderma pigmentosum donors in the comet assay: A potential diagnostic technique,Mutation Research, DNA Repair, 1992,273:137-144
22. Narendra P Singh et al, Abundant Alkali-Sensitive Site in DNA of Human and Mouse Sperm,Experimental Cell Research, 1989, 184:461-470
23. Narendra P Singh et al, A microgel electrophoresis technique for the direct quantitation of DNA damage and repair in individual fibroblasts cultured on microscope slides, Mutation Research ,1991,252:289-296
24. Olive P.L., et al, Heterogeneity in radiation-induced DNA damage and repair in tumor and normal cells measured using the "comet" assay, Radiat. Res., 1990b, 122:86-94.
25. Olive P.L., et al, Predicting tumor response to radiotherapy using the comet assay. Proceedings of the 4th International Conference on Time, Dose and Fractionation, in: B.R. Paliwal, T.J. Kinsella, D Herbert and Jfowler (Eds) Radiation Oncology The American Institute of Physics, Woodbury, NY,pp. 1993d, 65-76.
26. Olive P.L., et al, Growth fraction of human tumor: assay and complication. In/; R.E. Durand (ED.),Seminars in Radiation Oncology, Vol. 7: Cell Kinetics/: Application to Cancer Therapy, 1993e,3:90-98.
27. Olive P.L, et al, Induction and rejoining of radiation-induced DNA single strand breaks: "tail moment" as a function of position in the cell cycle. Mutat. Res., 1993b, 294:275-283.
28. Olive P.L, et al, Factors in Influencing DNA Migration from Individual Cells Subjected to Gel Electrophoresis. Exp.cell Res., 1992b, 198:259-267
29. Olive, P.L., Cell proliferation as a requirement for development of the contact effect in Chinese hamster V79 spheroids,Radiat. Res., 1989,117:79-92
30. Olive, P.L., J.P. Banath and C.D. Fjell, Influence of DNA strand breakage and DNA structure on staining by propidium iodide using the comet assay, Cytometry, in press, 1994a
31. Olive P.L, et al, DNA double strand breakes measured in individual cells subjected to gel electrophoresis. Cancer Res., 1991,51:4671
32. Ostling O.and K.J. Johanson, Microelectrophoretic study of radiation-induced DNA damages in individual mammalian cells, Biochemical and Biophysical Research commumition, 1984, 123(1):291-298.
33. Pooi-zobel BL, et al, Detection of genotoxicity in cells of the gastrointestinal tract. Mutagenesis,1992,7:166
34. Pool-Zobek et al, Systemic genotoxic effects of tobacco-related nitrosamines following oral and inhalation administration to Sprague-Dawley rats, Clin. Invest., 1992a, 70:299-306
35. Rojas E, et al, DNA damage in exfoliated buccal cells of smokes assessed by the single cell gel electrophoresis assay. Murat. Res., 1996,370:115-120.
36. Rydberg. B and K.J. Johanson, Estimation of DNA strand breaks in single mammalian cells, in: P.C.Hanawait, E.C. Friedberg and C.F. Fox (Eds.), DNA Repair Mechanisms,Academic Press, New York, 1978, pp465-468
37. Rydberg. B, Detection of induced DNA strand breaks with improved sensitivity in human cells,Radiat. Res, 1980,81:492-495.
38. Santos SJ, et al, Fluorescence in Situ Hybridization with Comets. Exp.cell Res.,1997,232(2):407-411.
39. Sardas S, et al, Assessment of smoking-induced DNA damage in iymphocytes of smoking mothers of newborn infants using the alkaline single cell gel electrophoresis technique. Mutat. Res.,1995,335:213-217.
40. Sardas S, et al, Assessment of DNA strand breakage by comet assay in diabetic patients and the role of antioxidant supplementation. Mutat. Res., 2001,490(2): 123-129.
41. Sasaki YF, et al, In vivo gentoxicity of ortho phenylphenol,biphenyl,and thiabendazole detected in multiple mouse organs by the alkaline single cell gel electrophoresis assay.Mutat. Res.,1997,395:189-198.
42. Singh NP et al, DNA damage and repair with age in individual human lymphocytes, Mutation Research, 1990, 237:123-130
43. Singh NP et al, A Simple Technique for Quantitation of Low Levels of DNA Damage in individual Cells, Experimental Cell Research, 1988,175:184-191
44. Singh NP et al, Modifications of alkaline microgel electrophoresis for sensitive detection of DNA damage, Int.J Radiat. Biol.,1994, 66:23-28
45. Singh NP, et al, A microgel electrophoresis technique for the direct quantitation od DNA gamage and repair in individual fibroblasts cultured on microscope slide. Mutat. Res., 1991,252:289
46. Singh NP, et al, Abundant alkali sensitive sites in DNA of human and mouse sperm. Expel. Cell Res., 1989, 184:461
47. Stavrova, et al, Single cell gel electrophoresis analysis of genomic damage induced by ethyl methanesuifonate in cultured tobacco cells. Mutat. Res.,1998,422(2):323-330.
48. Sutherland BM, et al, DNA Damage and Repair in Human Tissues. Nem York:Plenum press,1990,291-301
49. Tuck A, et al, Chornic lymphocytic leukemia lymphocytes lack the capacity to repair UVC-induced lesions. Mutat. Res., 2000,459(1):73-80
50. Tuo JS, et al, Benzene-induced genotoxicity in mice in vivo detected by the alkaline comet assay:reduction by CYPZEI inhibition. Murat. Res., 1996,368:213-219.
51. Udumudi A, et al,assesstion in cervical dyspasia patients by single cell gel electrophoresis assay: a study of DNA damage and repair. Mutat. Res., 1998,412:195-205.
52. Vijayalaxmi, Gary H.S. Strauss, and Raymond R.Tice, An analysis of γ-ray induced DNA
damage in human blood ieukocytes lymphocytes and granuloytes, Mutation Research,1993,292:123-128
53. Vijayalaxmi, Raymond R.Tice and Gary H.S. Strauss, Assessment of radiation-induced DNA damage in human blood lymphocytes using the single-ceil gel eletrophoresis technique, Mutation Research, 1992,271:243-252
54. Vijaya Lakshmi AN, et al, Detection of influenza virus induced DNA damage by comet assay.Mutat. Res., 1999,442:53-58,
55. McKelvey-Martin V.J.et al, The single cell gel electrophoresis assay(comet assay): A European review, Mutation Research,1993,288:47-63
56. Whitaker, S.J., S.N. powell and T.J. McMillan, Molecular assay of radiation-induced DNA damage,Eur.J Cancer,1991, 27:922-928
57. Yu Fs. et al, Detedtion of chemically induced DNA lesions in multiple mouse organs(liver, lung,spleen, kidney, and bone marrow) using the alkaline single cellgel electrophoresis (comet) assay.Murat. Res., 1997,373:31-37.
58. Zhang H,et al, Reduction of tumor hypoxia and inhibition of DNA repair by nicotinamide after irradiation of SCCV Ⅱ murine tumors and normal tissues.Cancer Res., 1996,56:2801 2808.
59．陈安，单细胞凝胶电泳法检测单细胞DNA损伤，国外医学临床生物化学与检测学分册，1998，19(4)：145
60．顾勤明 汪雪生 徐燕英，γ-射线对淋巴细胞DNA的损伤，《职业与健康》，2003，19(10)：3—5
61．韩宗超 张苏明，单细胞凝胶电泳在DNA损伤与修复检测中的应用，中华检验医学杂志，2000，23(3)：180
62．何惧 刘玉清，单细胞凝胶电泳技术的研究进展与应用，国外医学卫生学册，1997，24(2)：85
63．黄晓兰 杨明亮 吴晓 闫俊 罗琼，枸杞多糖对小鼠睾丸细胞DNA损伤的保护作用，卫生研究，2003，32(6)：599—561
64．李达圣 Kanehisa Morimoto Tatsuya Takeshita 吕玉泉，砷对人淋巴细胞DNA氧化损伤的作用，中华预防医学杂志，2002，36(1)：12—15
65．刘晓梅 石龙 金明华 王华 张哲雄 孙志伟 铅和镉对雄性小鼠生殖细胞DNA损伤的研究，中国公共卫生，2004，20(3)：311—312
66．刘轩军 齐晓东 范来富 徐辉，三氯苯诱发小鼠淋巴细胞DNA损伤的实验研究，中国工业医学杂志，2003，16(1)：32—34
67．孟紫强 张连珍，应用SCGE技术研究细胞DNA损伤的原理与方法，中国生物化学与分子生物学报，1998，14(5)：604—608
68．彭宝成 吴淑芬，单细胞凝胶电泳分析(彗星实验)及应用，河北医科大学学报，1997，18(4)：247
69．王芳 张双全 戴祝英，抗菌肽c M4组分对K562癌细胞染色质DNA断裂作用的SCGE研究，生物化学与生物物理进展，1998，25(1)：64—67
70．王海燕 张连珍 池翠萍，用单细胞微凝胶电泳技术(SCGE)检测电离辐射诱导三种细胞DNA损伤的研究，山西医药杂志，1999，28(5)：397—399
71．王玉春 赵荣材等，喹烯酮对小白鼠的致癌试验，中国兽医科技，1995，25(3)：24—25
72．谢麟，喹乙醇的安全性毒理学评价与合理应用，兽药与饲料添加剂，1999，5：25—27
73．邢彩虹 李桂兰 尹松年，单细胞凝胶电泳试验在人群生物监测研究中的应用审校，国外医学卫生学分册，2003，30(6)：377—382
74．许言午 张江虹 杨文敏，碱性单细胞凝胶电泳技术，山西医科大学学报，2003,34(2)：168—169
75．严相林 李金善 王玉春 赵荣材 薛飞群，喹烯酮的Ames试验，中兽医医药杂志，1998，5：11—12
76．严相林 李金善 王玉春 赵荣材 薛飞群，“喹烯酮”对小白鼠精子的畸变试验，中兽医医药杂志，1997，5：13—14
77．杨锦蓉 王菁 孙霆，CS_2诱导人淋巴细胞DNA损伤的再研究，浙江预防医学，2004，16(1)：1—3
78．姚洪菊 卜黎明，单细胞凝胶电泳检测吸烟者外周血淋巴细胞DNA的损伤，中国地方病学杂志，2002，21(5)：404—406
79．张爱华 黄晓欣 李军 蒋宪瑶 杨朝林，用SCGE法检测燃煤污染型砷中毒患者血细胞DNA的损伤作用，中国地方病学杂志，2000，19(1)：7—9
80．张波 刘承芸 孟紫强，单细胞凝胶电泳技术在环境污染物检测中的应用，环境科学与技术，2003，26(4)：42—45
81．张建平，单细胞凝胶电泳技术及其应用，国外医学遗传学分册，1997，20(5)：231
82．张全凤，喹乙醇的毒副作用及临床应用，福建畜牧兽医，2001，3(23)：29
83．张遵真等，单细胞凝胶电泳试验的最适条件研究，卫生毒理学杂志，1998，12(4)：249
84．朱靳良 邓丽霞 李芳红 蔡颖 郑履康 庄志雄，用单细胞凝胶电泳检测砷、汞、铅致DNA损伤的研究，卫生毒理学杂志，1998， 12(3)：141—143
85．庄志雄，细胞与分子生物学新技术在我国毒理学研究中的应用，卫生毒理学杂志，1999，13(4)：239
86．马爱国Collins AR等，不同细胞DNA损伤及自身修复能力的分析，癌变畸变突变，1997，9：138—142
87．杨丽 徐镜波 刘征涛，苯酚类化合物致突变Ames试验研究，东北师大学报自然科学版，2003，35(1)：82—85
2. Betti C,Banath TP, Durand RE, Comparative studies by Comet test and SCGE analysis in human lymphocytes from 200 healthy subjects, [J]. Mutation Research,1994,306:323-333.
3. Betti C, DaviniT, GiannessiL, et al, Comparative studies by Comet test and SCGE analysis in human lymphocytes from 200 healthy subjects, [J]. Mutation Research, 1995,343:201-207.
4. Betti C, et al, Microgel electrophoresis assay (comet test) and SCG analysis in human lymphocytes from 100 normal subjects. Mutat. Res., 1994,307:323-333.
5. Cerutli P.A., Prooxidant states and tumor promotion. Science,1985,227:375-381.
6. Anderson D.et al, The effect of various antioxidants and other modifying agents on oxygen-radical-generated DNA damage in human lymphocytes in the COMET assay, Mutation Research, 1994,307:261-271
7. Daryl W. Fairbairn et al,The cometassay:a comprehensive review, Mutation Research,1995,339:37-59
8. De Meo, M. Laget et al, Genotoxic activity of potassium permangannate in acidic solution,Mutation Research,1991,260:295-306
9. Duthie SJ, et al, The effect of dietary flavonoida on DNA damage (strand breaks and oxidized pyrimdines) and growth in human cells. Mutat. Res.,1997,390:141-151.
10. Gedik CM, et ai, Single cell electrophoresis applied to the analysis of UV-C damage and its repair in human cells, Int. J Radiat. Biol, 1992,62:313
11. Gellik, C.M., S.W.B. Ewen and A.R. Collins, Single-cell gel electrophoresis applied to the analysis of UV-C damage and its repair in human cells, Int.J Radiat. Biol.,1992, 62:313-320
12. Green, M.H.L. et ai, UV-C sensitivity of unstimulated and stimulated human lymphocytes from normal and xeroderma pigmentosum donors in the comet assay: A potential diagnostic technique,Mutation Research, 1992, 273:137-144
13. Grichuer T, et al, Induction of somatic DNA damage as measure by single cell gel electrophoresis and point mutation in leaves of tobacco plants. Mutat. Res., 1998,401 (2):143-152
14. Gutierrez S,et al,Application of the single tee gel electrophoresis(SCGE) assaytothe detected of DNA damage induced by 1311 treatment in hyperthyroidism patients. Mutagenesis, 1998,13:95 98.
15. Guyton, et al, Oxidative mechanisms in carcinogenesis, Br. Med. Bull., 1993, 49:523-544
16. Heliman B, et al, the conpects of tail moment and tail inertia in the single cell gel electrophoresis assay. Murat. Res., 1995,336:123-131
17. Lubin JH, Blot WJ, Berrino FM, odifying risk of developing lung cancer by changing habits of cigarette smoking, [J].BrMedJ, 1984,288:1953-1956
18. Maarit HL, et al, Repair of γ-irradiation-induced DNA single-strand breaks in human
bonemarrow cells: Effects of a second irradiation. Mutat. Res., 1997, 388:33-44.
19. Maes A, et al, Cytogenetic effects of 935-2 MH_2 (GSM) microwaves a lone and in combition with mitomycinC, Mutat. Res., 1997,393(1-2):151
20. Malyapa R A, et al, DNA damage in rat brain cells after in vivo exposure to 2450 MH2 electromagnetic radiation and various methods of euthanasia. Radiat Res.,1998,149(6):637-645
21..Green M.H.L et al, UV-C sensitivity of unstimulated and stimulated human lymphocytes from normal and xeroderma pigmentosum donors in the comet assay: A potential diagnostic technique,Mutation Research, DNA Repair, 1992,273:137-144
22. Narendra P Singh et al, Abundant Alkali-Sensitive Site in DNA of Human and Mouse Sperm,Experimental Cell Research, 1989, 184:461-470
23. Narendra P Singh et al, A microgel electrophoresis technique for the direct quantitation of DNA damage and repair in individual fibroblasts cultured on microscope slides, Mutation Research ,1991,252:289-296
24. Olive P.L., et al, Heterogeneity in radiation-induced DNA damage and repair in tumor and normal cells measured using the "comet" assay, Radiat. Res., 1990b, 122:86-94.
25. Olive P.L., et al, Predicting tumor response to radiotherapy using the comet assay. Proceedings of the 4th International Conference on Time, Dose and Fractionation, in: B.R. Paliwal, T.J. Kinsella, D Herbert and Jfowler (Eds) Radiation Oncology The American Institute of Physics, Woodbury, NY,pp. 1993d, 65-76.
26. Olive P.L., et al, Growth fraction of human tumor: assay and complication. In/; R.E. Durand (ED.),Seminars in Radiation Oncology, Vol. 7: Cell Kinetics/: Application to Cancer Therapy, 1993e,3:90-98.
27. Olive P.L, et al, Induction and rejoining of radiation-induced DNA single strand breaks: "tail moment" as a function of position in the cell cycle. Mutat. Res., 1993b, 294:275-283.
28. Olive P.L, et al, Factors in Influencing DNA Migration from Individual Cells Subjected to Gel Electrophoresis. Exp.cell Res., 1992b, 198:259-267
29. Olive, P.L., Cell proliferation as a requirement for development of the contact effect in Chinese hamster V79 spheroids,Radiat. Res., 1989,117:79-92
30. Olive, P.L., J.P. Banath and C.D. Fjell, Influence of DNA strand breakage and DNA structure on staining by propidium iodide using the comet assay, Cytometry, in press, 1994a
31. Olive P.L, et al, DNA double strand breakes measured in individual cells subjected to gel electrophoresis. Cancer Res., 1991,51:4671
32. Ostling O.and K.J. Johanson, Microelectrophoretic study of radiation-induced DNA damages in individual mammalian cells, Biochemical and Biophysical Research commumition, 1984, 123(1):291-298.
33. Pooi-zobel BL, et al, Detection of genotoxicity in cells of the gastrointestinal tract. Mutagenesis,1992,7:166
34. Pool-Zobek et al, Systemic genotoxic effects of tobacco-related nitrosamines following oral and inhalation administration to Sprague-Dawley rats, Clin. Invest., 1992a, 70:299-306
35. Rojas E, et al, DNA damage in exfoliated buccal cells of smokes assessed by the single cell gel electrophoresis assay. Murat. Res., 1996,370:115-120.
36. Rydberg. B and K.J. Johanson, Estimation of DNA strand breaks in single mammalian cells, in: P.C.Hanawait, E.C. Friedberg and C.F. Fox (Eds.), DNA Repair Mechanisms,Academic Press, New York, 1978, pp465-468
37. Rydberg. B, Detection of induced DNA strand breaks with improved sensitivity in human cells,Radiat. Res, 1980,81:492-495.
38. Santos SJ, et al, Fluorescence in Situ Hybridization with Comets. Exp.cell Res.,1997,232(2):407-411.
39. Sardas S, et al, Assessment of smoking-induced DNA damage in iymphocytes of smoking mothers of newborn infants using the alkaline single cell gel electrophoresis technique. Mutat. Res.,1995,335:213-217.
40. Sardas S, et al, Assessment of DNA strand breakage by comet assay in diabetic patients and the role of antioxidant supplementation. Mutat. Res., 2001,490(2): 123-129.
41. Sasaki YF, et al, In vivo gentoxicity of ortho phenylphenol,biphenyl,and thiabendazole detected in multiple mouse organs by the alkaline single cell gel electrophoresis assay.Mutat. Res.,1997,395:189-198.
42. Singh NP et al, DNA damage and repair with age in individual human lymphocytes, Mutation Research, 1990, 237:123-130
43. Singh NP et al, A Simple Technique for Quantitation of Low Levels of DNA Damage in individual Cells, Experimental Cell Research, 1988,175:184-191
44. Singh NP et al, Modifications of alkaline microgel electrophoresis for sensitive detection of DNA damage, Int.J Radiat. Biol.,1994, 66:23-28
45. Singh NP, et al, A microgel electrophoresis technique for the direct quantitation od DNA gamage and repair in individual fibroblasts cultured on microscope slide. Mutat. Res., 1991,252:289
46. Singh NP, et al, Abundant alkali sensitive sites in DNA of human and mouse sperm. Expel. Cell Res., 1989, 184:461
47. Stavrova, et al, Single cell gel electrophoresis analysis of genomic damage induced by ethyl methanesuifonate in cultured tobacco cells. Mutat. Res.,1998,422(2):323-330.
48. Sutherland BM, et al, DNA Damage and Repair in Human Tissues. Nem York:Plenum press,1990,291-301
49. Tuck A, et al, Chornic lymphocytic leukemia lymphocytes lack the capacity to repair UVC-induced lesions. Mutat. Res., 2000,459(1):73-80
50. Tuo JS, et al, Benzene-induced genotoxicity in mice in vivo detected by the alkaline comet assay:reduction by CYPZEI inhibition. Murat. Res., 1996,368:213-219.
51. Udumudi A, et al,assesstion in cervical dyspasia patients by single cell gel electrophoresis assay: a study of DNA damage and repair. Mutat. Res., 1998,412:195-205.
52. Vijayalaxmi, Gary H.S. Strauss, and Raymond R.Tice, An analysis of γ-ray induced DNA
damage in human blood ieukocytes lymphocytes and granuloytes, Mutation Research,1993,292:123-128
53. Vijayalaxmi, Raymond R.Tice and Gary H.S. Strauss, Assessment of radiation-induced DNA damage in human blood lymphocytes using the single-ceil gel eletrophoresis technique, Mutation Research, 1992,271:243-252
54. Vijaya Lakshmi AN, et al, Detection of influenza virus induced DNA damage by comet assay.Mutat. Res., 1999,442:53-58,
55. McKelvey-Martin V.J.et al, The single cell gel electrophoresis assay(comet assay): A European review, Mutation Research,1993,288:47-63
56. Whitaker, S.J., S.N. powell and T.J. McMillan, Molecular assay of radiation-induced DNA damage,Eur.J Cancer,1991, 27:922-928
57. Yu Fs. et al, Detedtion of chemically induced DNA lesions in multiple mouse organs(liver, lung,spleen, kidney, and bone marrow) using the alkaline single cellgel electrophoresis (comet) assay.Murat. Res., 1997,373:31-37.
58. Zhang H,et al, Reduction of tumor hypoxia and inhibition of DNA repair by nicotinamide after irradiation of SCCV Ⅱ murine tumors and normal tissues.Cancer Res., 1996,56:2801 2808.
59．陈安，单细胞凝胶电泳法检测单细胞DNA损伤，国外医学临床生物化学与检测学分册，1998，19(4)：145
60．顾勤明 汪雪生 徐燕英，γ-射线对淋巴细胞DNA的损伤，《职业与健康》，2003，19(10)：3—5
61．韩宗超 张苏明，单细胞凝胶电泳在DNA损伤与修复检测中的应用，中华检验医学杂志，2000，23(3)：180
62．何惧 刘玉清，单细胞凝胶电泳技术的研究进展与应用，国外医学卫生学册，1997，24(2)：85
63．黄晓兰 杨明亮 吴晓 闫俊 罗琼，枸杞多糖对小鼠睾丸细胞DNA损伤的保护作用，卫生研究，2003，32(6)：599—561
64．李达圣 Kanehisa Morimoto Tatsuya Takeshita 吕玉泉，砷对人淋巴细胞DNA氧化损伤的作用，中华预防医学杂志，2002，36(1)：12—15
65．刘晓梅 石龙 金明华 王华 张哲雄 孙志伟 铅和镉对雄性小鼠生殖细胞DNA损伤的研究，中国公共卫生，2004，20(3)：311—312
66．刘轩军 齐晓东 范来富 徐辉，三氯苯诱发小鼠淋巴细胞DNA损伤的实验研究，中国工业医学杂志，2003，16(1)：32—34
67．孟紫强 张连珍，应用SCGE技术研究细胞DNA损伤的原理与方法，中国生物化学与分子生物学报，1998，14(5)：604—608
68．彭宝成 吴淑芬，单细胞凝胶电泳分析(彗星实验)及应用，河北医科大学学报，1997，18(4)：247
69．王芳 张双全 戴祝英，抗菌肽c M4组分对K562癌细胞染色质DNA断裂作用的SCGE研究，生物化学与生物物理进展，1998，25(1)：64—67
70．王海燕 张连珍 池翠萍，用单细胞微凝胶电泳技术(SCGE)检测电离辐射诱导三种细胞DNA损伤的研究，山西医药杂志，1999，28(5)：397—399
71．王玉春 赵荣材等，喹烯酮对小白鼠的致癌试验，中国兽医科技，1995，25(3)：24—25
72．谢麟，喹乙醇的安全性毒理学评价与合理应用，兽药与饲料添加剂，1999，5：25—27
73．邢彩虹 李桂兰 尹松年，单细胞凝胶电泳试验在人群生物监测研究中的应用审校，国外医学卫生学分册，2003，30(6)：377—382
74．许言午 张江虹 杨文敏，碱性单细胞凝胶电泳技术，山西医科大学学报，2003,34(2)：168—169
75．严相林 李金善 王玉春 赵荣材 薛飞群，喹烯酮的Ames试验，中兽医医药杂志，1998，5：11—12
76．严相林 李金善 王玉春 赵荣材 薛飞群，“喹烯酮”对小白鼠精子的畸变试验，中兽医医药杂志，1997，5：13—14
77．杨锦蓉 王菁 孙霆，CS_2诱导人淋巴细胞DNA损伤的再研究，浙江预防医学，2004，16(1)：1—3
78．姚洪菊 卜黎明，单细胞凝胶电泳检测吸烟者外周血淋巴细胞DNA的损伤，中国地方病学杂志，2002，21(5)：404—406
79．张爱华 黄晓欣 李军 蒋宪瑶 杨朝林，用SCGE法检测燃煤污染型砷中毒患者血细胞DNA的损伤作用，中国地方病学杂志，2000，19(1)：7—9
80．张波 刘承芸 孟紫强，单细胞凝胶电泳技术在环境污染物检测中的应用，环境科学与技术，2003，26(4)：42—45
81．张建平，单细胞凝胶电泳技术及其应用，国外医学遗传学分册，1997，20(5)：231
82．张全凤，喹乙醇的毒副作用及临床应用，福建畜牧兽医，2001，3(23)：29
83．张遵真等，单细胞凝胶电泳试验的最适条件研究，卫生毒理学杂志，1998，12(4)：249
84．朱靳良 邓丽霞 李芳红 蔡颖 郑履康 庄志雄，用单细胞凝胶电泳检测砷、汞、铅致DNA损伤的研究，卫生毒理学杂志，1998， 12(3)：141—143
85．庄志雄，细胞与分子生物学新技术在我国毒理学研究中的应用，卫生毒理学杂志，1999，13(4)：239
86．马爱国Collins AR等，不同细胞DNA损伤及自身修复能力的分析，癌变畸变突变，1997，9：138—142
87．杨丽 徐镜波 刘征涛，苯酚类化合物致突变Ames试验研究，东北师大学报自然科学版，2003，35(1)：82—85
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