降低酶联核酸适配体分析方法中非特异性吸附的分析研究

以检测蜂蜜中的四环素残留为例，构建间接竞争酶联核酸适体分析法（ic-ELAA），就ic-ELAA中非特异性吸附的影响因素进行分析，探讨降低非特异性吸附的措施。结果表明：选择2 μg/mL四环素-牛血清蛋白在10 mmol/L Tris-HCl（pH 8.0）中过夜包被，次日用0.5 g/mL酪蛋白封闭1 h，竞争反应后用6 μg/mL链霉亲和素-辣根过氧化物酶催化颜色反应放大竞争效果，最后采用450 nm和630 nm双波长读数，可得到灵敏度为0.035 ng/mL的竞争曲线。同时，根据长期实验经验，提出其他降低非特异性吸附的注意事项。

Methods of reducing non-specific adsorption in an indirect competitive enzyme-linked aptamer assay

An indirect competitive enzyme-linked aptamer assay for tetracycline detection in honey has been developed. We analyzed different parameters which may cause non-specific adsorption in the detection system and employed different methods to reduce the non-specific adsorption. According to the detailed optimization, 2 μg/mL of TC-BSA was diluted in 10 mmol/L Tris-HCl (pH 8.0) and incubated in microplates overnight, and then the wells were blocked with 0.5 g/mL casein for 1 h, followed by the competitive reaction. The competition was then amplified by 6 μg/mL of SA-HRP and the final absorbance signals were collected with a double-wavelength spectrometer. Under the optimized conditions, this indirect competitive enzyme-linked aptamer assay can be carried out to detect tetracycline in honey samples, with a limit of detection of 0.035 ng/mL. In addition, several factors ensuring the good performance of the enzyme-linked aptamer assay are highlighted.