Nelson N C, Cheikh A B, Matsuda E, Becker M M
Gen-Probe Incorporated, San Diego, California 92121, USA.
Biochemistry. 1996 Jun 25;35(25):8429-38. doi: 10.1021/bi960085+.
The acridinium ester 4-(2-succinimidyloxycarbonylethyl)phenyl-10-methylacridinium 9-carboxylate trifluoromethane sulfonate (AE), which reacts rapidly with alkaline hydrogen peroxide to produce light, has been used as a detection label in a number of assay procedures, including nucleic acid probe-based systems [Nelson et al. (1995) in Nonisotopic Probing, Blotting and Sequencing (Kricka, L. J., Ed.) pp 391-428, Academic Press, Inc., San Diego, CA]. We have synthesized a number of derivatives of this AE and characterized their chemiluminescent properties. These derivatives display significant differences in the kinetics of the chemiluminescence reaction as well as optimal pH for light production. These differences allow two or more derivatives to be simultaneously detected and quantitated in a single reaction vessel. Several of these derivatives have been covalently linked to nucleic acid probe molecules and have been further characterized in regard to chemiluminescence properties as well as hydrolysis of the ester bond in both single- and double-stranded conformations. On the basis of these properties, homogeneous assay formats utilizing DNA probes labeled with various AE derivatives were developed. Simultaneous detection and quantitation of Chlamydia trachomatis and Neisseria gonorrhoeae, the gag and pol regions of HIV, and wild-type and mutant HIV sequences was achieved with high sensitivity and discrimination.
吖啶酯4-(2-琥珀酰亚胺氧基羰基乙基)苯基-10-甲基吖啶鎓9-羧酸三氟甲磺酸盐（AE）能与碱性过氧化氢快速反应产生光，已在包括基于核酸探针的系统等多种检测程序中用作检测标记物[纳尔逊等人（1995年），载于《非同位素探测、印迹和测序》（克里卡，L. J.编），第391 - 428页，学术出版社，加利福尼亚州圣地亚哥]。我们合成了该AE的多种衍生物，并对其化学发光性质进行了表征。这些衍生物在化学发光反应动力学以及发光的最佳pH值方面表现出显著差异。这些差异使得两种或更多种衍生物能够在单个反应容器中同时被检测和定量。其中几种衍生物已与核酸探针分子共价连接，并在单链和双链构象下就化学发光性质以及酯键水解情况进行了进一步表征。基于这些性质，开发了利用用各种AE衍生物标记的DNA探针的均相检测方法。实现了对沙眼衣原体和淋病奈瑟菌、HIV的gag和pol区域以及野生型和突变型HIV序列的同时高灵敏度检测和定量。
