5-甲基-2'-脱氧胞苷的时间分辨荧光免疫分析

Time-resolved fluoroimmunoassay of 5-methyl-2'-deoxycytidine.

作者信息

Vilpo J A, Rasi S, Suvanto E, Vilpo L M

出版信息

Anal Biochem. 1986 May 1;154(2):436-40. doi: 10.1016/0003-2697(86)90011-4.

Abstract

We describe time-resolved fluoroimmunoassay of 5-methyl-2'-deoxycytidine (5MedCyd). The assay is based on the use of a highly specific antiserum raised in rabbits against BSA-conjugated 5-methylcytidine (5MeCyd). The tracer in the solid-phase time-resolved fluoroimmunoassay (TR-FIA) was antigen-selected anti-5MedCyd labeled with Europium. Thyroglobulin-linked 5MeCyd served as the solid-phase antigen. The measuring range for the fluoroimmunoassay was from less than 1 to 5000 pmol per assay of 5MedCyd. A good correlation between the results obtained with the TR-FIA and HPLC was demonstrated when the methods were applied to the measurement of methylation in human leukemic cells and other DNA samples. TR-FIA has several advantages over the more laborious techniques available so far: (i) high sensitivity, (ii) large assay ranges, (iii) rapidity and large number of simultaneous assays, (iv) simplicity, and (v) low cost provided that the laboratory has equipment for time-resolved fluorometry.

摘要

我们描述了5-甲基-2'-脱氧胞苷（5MedCyd）的时间分辨荧光免疫测定法。该测定法基于使用在兔体内产生的针对牛血清白蛋白偶联的5-甲基胞苷（5MeCyd）的高度特异性抗血清。固相时间分辨荧光免疫测定法（TR-FIA）中的示踪剂是用铕标记的抗原选择的抗5MedCyd。甲状腺球蛋白连接的5MeCyd用作固相抗原。荧光免疫测定法的测量范围为每次测定5MedCyd小于1至5000皮摩尔。当将这些方法应用于人类白血病细胞和其他DNA样品的甲基化测量时，TR-FIA获得的结果与高效液相色谱法之间显示出良好的相关性。与目前可用的更繁琐的技术相比，TR-FIA具有几个优点：（i）高灵敏度，（ii）大测定范围，（iii）快速性和大量同时测定，（iv）简单性，以及（v）如果实验室拥有时间分辨荧光测定设备，则成本低。