Cross-linking of cytokeratins to DNA in vivo by chromium salt and cis-diamminedichloroplatinum(II).
Cross-linking of cytokeratins to DNA in vivo by chromium salt and cis-diamminedichloroplatinum(II).
复制标题
通过铬盐和顺式二氯二氨铂 (II) 在体内将细胞角蛋白与 DNA 交联。
DOI：
10.1021/bi00349a001
复制
发表时间：
1986
期刊：
影响因子：
2.9
通讯作者：
Hnilica,LS
中科院分区：
文献类型：
--
作者：
Wedrychowski,A;Schmidt,WN;Ward,WS;Hnilica,LS
材料和方法材料。 m-DDP 购自 Sigma Chemical Co.（密苏里州圣路易斯）。 K2Cr04 来自 Fisher Scientific Co.（宾夕法尼亚州匹兹堡）。 DNase I 和 RNase A 来自 Worthington Biochemical Corp.（弗里霍尔德，新泽西州）。电泳用品来自 Bio-Rad Laboratories（里士满，加利福尼亚州）。每 5 天将 Novikoff 腹水肝癌 (NAH) 细胞传给雄性 Sprague-Dawley 大鼠（Harlan Industries，印第安纳波利斯，印第安纳州）。收获细胞，重悬于冰冷的 0.17 Mnh4c1 中以裂解污染的红细胞，在 PBS 中洗涤，并在交联实验中新鲜使用。交联实验。采用相同的程序与m-DDP和K 2 CrO 4 进行交联。将洗涤后的 NAH 细胞重悬于 20 体积（包装细胞/缓冲液，v/v）的含有不同浓度交联剂的 Hank 平衡盐溶液中，并在 37°C 下孵育指定的时间间隔。孵育期间，每 20 分钟轻轻摇动细胞一次。 Hank 平衡溶液除了含有等渗浓度的 NaCl 和 cf-葡萄糖外，还含有大量的 Ca2+、Mg2+ 和 K+ 离子。由于它们的解离状态，这些阳离子可以与培养介质中的铬阴离子发生反应。他们的反应
Materials and MethodsMaterials. m-DDP was purchased from Sigma Chemical Co.(St. Louis, MO). K2Cr04 was from Fisher Scientific Co.(Pittsburgh, PA). DNase I and RNase A were from Worthington Biochemical Corp.(Freehold, NJ). Electro-phoresis supplies were from Bio-Rad Laboratories (Richmond, CA).Novikoff ascites hepatoma (NAH) cells were passed in male Sprague-Dawley rats (Harlan Industries, Indianapolis, IN) every 5 days. Cells were harvested, resuspended in ice-cold 0.17 Mnh4c1 to lyse contaminating erythrocytes, washed in PBS, and used freshly in the cross-linking experiments. Cross-Linking Experiments. Identical procedures were employed for cross-linking with m-DDP and K2Cr04. The washed NAH cells were resuspended in 20 volumes (packed cell/buffer, v/v) of Hank’s balanced salt solution containing different concentrations of cross-linking agent and incubated at 37 C for indicated time intervals. During incubation the cells were gently shaken every 20 min. The Hank’s balanced solution contains, in addition to isotonic concentrations of NaCl and cf-glucose, significant amounts of Ca2+, Mg2+, and K+ ions. Because of their dissociated states, these cations can react with chromium anions in the incubation medium. Their reaction