Reproducibility of human sperm DNA measurements using the alkaline single cell gel electrophoresis assay

被引：94

作者：

Hughes, CM ^{[1
]}

Lewis, SEM ^{[1
]}

McKelveyMartin, VJ ^{[1
]}

Thompson, W ^{[1
]}

机构：

[1] UNIV ULSTER,CANC & AGEING RES GRP,SCH BIOMED SCI,COLERAINE BT52 1SA,LONDONDERRY,NORTH IRELAND

来源：

MUTATION RESEARCH-FUNDAMENTAL AND MOLECULAR MECHANISMS OF MUTAGENESIS | 1997年 / 374卷 / 02期

关键词：

human spermatozoon; comet assay; reproducibility; DNA damage; STRAND BREAKS; SPERMATOZOA; RADIATION; DAMAGE;

D O I：

10.1016/S0027-5107(96)00241-2

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

The single cell gel electrophoresis (SCGE) assay is a simple visual technique used to assess DNA integrity in individual cells by measuring damage reflected as strand breaks under alkaline conditions. Cells are embedded in agarose on glass slides followed by lysis of the cell membranes after which damaged DNA strands are electrophoresed away from the nucleus towards the anode and deposited to one side giving the appearance of a tail. DNA damage may be measured by assessing the relative amounts of DNA remaining in the head as opposed to those strands which have formed the tail. The assay has been used to determine DNA quality in human sperm (Hughes, C.M., S.E.M. Lewis, V.J. McKelvey-Martin, W. Thompson, A comparison of baseline and induced DNA damage in human sperm from fertile and infertile man, using a modified comet assay. Mol. Human Reprod., in press) by measuring fifty cells on one slide for each individual. Coefficients of variation between three control slides prepared for ten individuals were less than 4% and less than 9% for three slides prepared using irradiated sperm. Ten readings of fifty sperm each from a single slide showed a coefficient of variation of less than 6% for ten individuals studied. These results indicate that the measurement of fifty sperm from a single slide is sufficient to assess the DNA damage within a sperm population. Coefficients of variation of less than 5.4% for repeated analysis of individual cells were obtained which demonstrates the reproducibility of the image analysis software.

引用

页码：261 / 268

页数：8