睾酮—吖啶酯化学发光示踪剂的制备及评价

EVALUATION OF TESTOSTERONE-3-ACRIDINIUM ESTER CONJUGATE FOR CHEMILUMINESCENCE IMMUNOASSAY

本文报道了10-甲基-吖淀-9-核酸[4-(N一睾酮-3-羧甲氧基肟)-氯乙基]苯酯(To-3-AEPMAC)的合成、理化鉴定和免疫学评价.制备方法类似于以往甾体激素全抗原的合成,产品的化学发光信号与噪声比高,可测限为0.18fmol,线性可测范围可达6个数量级以上,与抗睾酮血清结合不改变吖啶酯的化学发光动力学和发光效率;免疫化学最高结合率和非特异结合稳定,抗血清稀释曲线比较证明产品的免疫反应性基本无损失,优于鲁米诺及其衍生物制备的示踪剂,符合建立超微量免疫化学分析中标记化合物的要求.

The effect of varying conditions to Testosterone-3-carboxymethyl oxime-4- (10-methy) -acridinium-9- carboxylate (To-3-AEPMAC) on the light yield and binding affinity of Testosterone chemiluminescent-marker conjugate was investigated. For this purpose To-3-AEPMAC has been synthesized, the resulting was then evaluted as potential labels for the development of an immunoassay based on monitoring chemiluminescence. The conjugate was able to compete with testosterone for the binding siles of an antibody raised against To-3-CMO bovine serum albumin, and they showed high affinity as same as unaltered testosterone. The conjugate produced light up on oxidation by a hydrogen peroxide in basic condition and was not significantly affected the quantum yield. The chemiluminescent reaction was optimized in terms of pH 6.0 concentration of oxidant, sodium hydrate and conjugate design. Under optimal conditions, the conjugate was detectable at femtomolar levels. The lowest detection limit was obtained using To-3-AEP-MAC (0.18fmol). These results indicated that conjugate does not modify its chemiluminescent property and immunoreativity,the immunoassay techniques based on chemiluminescence can be developed with the label.