Olive P L
British Columbia Cancer Research Centre, Vancouver, Canada.
Environ Mol Mutagen. 1988;11(4):487-95. doi: 10.1002/em.2850110409.
When mammalian cells are lysed in 2% sodium dodecyl sulphate detergent followed by addition of an equal volume of 0.12 M potassium chloride, a precipitate forms that can be collected by low-speed centrifugation. This precipitate contains the cell protein and nucleic acid in close association with protein. In the absence of DNA damage, most of the DNA precipitates, but when DNA strand breaks are created by exposing cells to ionizing radiation or toxic chemicals, DNA is released from the protein and remains in the supernatant after centrifugation. The proportion of DNA remaining in the supernatant is thus a measure of the amount of DNA damage. This assay is characterized in terms of optimum cell number and pH and dose-response curves for DNA damage and cell survival following ionizing radiation, MNNG, BCNU, and VP-16 are shown. Sensitivity, simplicity, speed, and large sample handling capacity should allow wide application of this new assay to a variety of questions concerning DNA damage and repair.
当哺乳动物细胞在2%十二烷基硫酸钠洗涤剂中裂解，随后加入等体积的0.12 M氯化钾时，会形成一种沉淀，可通过低速离心收集。这种沉淀包含与蛋白质紧密结合的细胞蛋白质和核酸。在没有DNA损伤的情况下，大部分DNA会沉淀下来，但当通过将细胞暴露于电离辐射或有毒化学物质而产生DNA链断裂时，DNA会从蛋白质中释放出来，并在离心后留在上清液中。因此，上清液中剩余DNA的比例是DNA损伤量的一种度量。该检测方法在最佳细胞数量、pH值以及电离辐射、MNNG、BCNU和VP - 16处理后DNA损伤和细胞存活的剂量反应曲线方面具有特征。灵敏度、简便性、速度和大样本处理能力应使这种新检测方法能够广泛应用于各种有关DNA损伤和修复的问题。
