基于非标记适配体结构变化荧光法检测黄曲霉毒素B1

A fluorescence method for detection of aflatoxin B1 based on structural change of unlabeled aptamer

构建了一种基于非标记适配体结构变化荧光检测黄曲霉毒素B1(AFB1)的方法。无AFB1时,一条非标记的AFB1适配体同时与2条短互补DNA链杂交,形成DNA双链结构,导致标记于其中一条互补DNA的3’端的荧光素(FAM)与标记于另一条互补DNA的5’端的淬灭剂(BHQ1)相邻近,发生荧光共振能量转移,FAM荧光被BHQ1淬灭。AFB1存在时,适配体与AFB1结合,而不与互补DNA发生杂交。此时,FAM与BHQ1距离较远,FAM荧光不能被淬灭。通过测量体系荧光强度变化可定量检测AFB1。方法检出限0.2 nmol/L,定量检测范围1.0 nmol/L~4.0μmol/L。该方法无需共价标记适配体,操作简便,特异性好,能够用于检测复杂基质样品中的AFB1。

A simple method was constructed for fluorescence detection of aflatoxin B1(AFB1)based on the structural change of an unlabeled aptamer.In the absence of AFB1,an unlabeled AFB1 aptamer simultaneously hybridized with two complementary short DNA strands,forming a DNA duplex structure.As results,the fluorescein FAM labeled at 3'end of a complementary DNA was adjacent to the quencher BHQ1 labeled at 5'end of the other complementary DNA.In this case,fluorescence resonance energy transfer occurred,and FAM fluorescence was quenched by BHQ1.In the presence of AFB1,the aptamer bound to AFB1 instead of hybridizing with complementary DNAs.As results,FAM and BHQ1 were far away from each other,and FAM fluorescence could not be quenched.AFB1 was quantitatively detected by measuring fluorescence intensity of the system.Detection limit of this method was 0.2 nmol/L,and quantitative detection range was 1.0 nmol/L-4.0μmol/L.This method could be used for rapid detection of AFB1 in complex matrix samples for not requiring labeling of aptamer,easy to operate,and good specificity.