Azek F, Grossiord C, Joannes M, Limoges B, Brossier P
Laboratoire de Microbiologie Médicale et Moléculaire, Faculté de Médecine et Pharmacie, 7 Boulevard Jeanned'Arc, Dijon, 21033, France.
Anal Biochem. 2000 Aug 15;284(1):107-13. doi: 10.1006/abio.2000.4692.
A disposable electrochemical biosensor for the detection of DNA sequences related to the human cytomegalovirus (HCMV) is described. The sensor relies on the adsorption of an amplified human cytomegalovirus DNA strand onto the sensing surface of a screen-printed carbon electrode, and to its hybridization to a complementary single-stranded biotinylated DNA probe. The extent of hybrids formed was determined with streptavidin conjugated to horseradish peroxidase. The peroxidase label was indirectly quantified by measuring the amount of the chromophore and electroactive product 2,2'-diaminoazobenzene generated from the o-phenylenediamine substrate. The intensity of differential pulse voltammetric peak currents resulting from the reduction of the enzyme-generated product was related to the number of target HCMV-amplified DNA molecules present in the sample, and the results were compared to those obtained with standard methods, i.e., agarose gel electrophoresis quantification and colorimetric hybridization assay in a microtiter plate. A detection limit of 0.6 amol/ml of HCMV-amplified DNA fragment was obtained with the electrochemical DNA biosensor. The electrochemical method was 23,000-fold more sensitive than the gel electrophoresis technique and 83-fold more sensitive than the colorimetric hybridization assay in a microtiter plate.
本文描述了一种用于检测与人类巨细胞病毒（HCMV）相关DNA序列的一次性电化学生物传感器。该传感器依靠将扩增的人类巨细胞病毒DNA链吸附到丝网印刷碳电极的传感表面，并使其与互补的单链生物素化DNA探针杂交。用与辣根过氧化物酶偶联的链霉亲和素测定形成的杂交体的程度。通过测量从邻苯二胺底物产生的发色团和电活性产物2,2'-二氨基偶氮苯的量来间接定量过氧化物酶标记。酶产生的产物还原所产生的差分脉冲伏安峰电流强度与样品中存在的目标HCMV扩增DNA分子数量相关，并将结果与用标准方法（即琼脂糖凝胶电泳定量和微量滴定板中的比色杂交测定）获得的结果进行比较。电化学生物传感器对HCMV扩增DNA片段的检测限为0.6 amol/ml。电化学方法比凝胶电泳技术灵敏23,000倍，比微量滴定板中的比色杂交测定灵敏83倍。
