用于检测牛奶中雌二醇的酶热传感技术研究

摘要

目的：建立一种用于检测牛奶中雌二醇的酶热传感技术。方法羧基化雌二醇经1-乙基-(3-二甲基氨基丙基)碳酰二亚胺盐酸盐(EDC)与β-内酰胺酶偶联。酶热传感器中装SPG酶柱,通过ELISA原理使游离雌二醇与结合雌二醇竞争结合雌二醇抗体,从而检测基质中的雌二醇浓度。优化酶热传感检测体系中的反应底物、系统流速、抗原抗体稀释比例等实验因素。结果磷酸盐缓冲液(PBS)基质中测定IC50值为10.2 ng/mL,线性检测范围：5.1~19.4 ng/mL,最低检测限2.8 ng/mL,变异系数为3.4%,与多种雌激素类结构类似物之间均无交叉反应,特异性良好。牛奶样品中测定IC50值：0.45 ng/mL,线性检测范围：0.24~0.79 ng/mL,最低检测限0.12 ng/mL。结论酶热传感技术可以快速检测牛奶中的雌二醇残留,操作便捷、样品前处理简单,是一种高效、经济的适用于食品安全工作现场检测的方法。%Objective To establish an enzyme thermistor technique for determination of estradiol in milk. Methods Carboxyl transformation and coupled estradiol withβ-lactamase through 1-ethyl-(3-dimethyl amino propyl) carbonyl imine hydrochloride (EDC). SPG was installed in thermal sensor as enzyme column, and direct competition ELISA principle was used to detect estradiol concentrations in matrix. Experimental factors such as the reaction substrate, the flow rate of the system, and the antigen-antibody dilution ratio were optimized.Results The linerity range was 5.1~19.4 ng/mL, IC50 was 10.2 ng/mL and the detection limit was 2.8 ng/mL in PBS sample. The result was stable, reliable and specific, no cross-reaction could be demonstrated with a variety of estrogen analogues and coefficient variation (CV) was 3.4%. The linerity range was 0.24~0.79 ng/mL, IC50was 0.45 ng/mL, and the detection limit was 0.12 ng/mL in milk sample.Conclusion This method is successfully applied to detect estradiol in milk, it is simple, efficient, economic, and suitable for the on-site use.