沙丁胺醇药物残留酶联免疫检测法的建立及优化

The Establishment and Optimization of Enzyme-linked Immunoassay Method for detection of Salbutamol Residue

本试验通过优化ELISA工作条件,建立间接竞争ELISA检测沙丁胺醇(SAL)的方法。采用方阵滴定法确定包被抗原、抗体、酶标二抗的工作浓度;以上述试验条件为基础,以系列稀释的SAL标准溶液作为检测对象,绘制标准曲线,并最终确定优化的试验方案。方阵滴定法确定包被抗原工作浓度为1:2000,抗体工作浓度为1:4000,酶标二抗工作浓度为1:6000;从标准曲线可知,线性检测范围为0.5～20 ng/mL,IC50为2.33 ng/mL;最终优化的试验方案中酶标板为深圳金灿华有限公司的产品,包被液为0.05 mol/LpH 9.6的碳酸盐缓冲液,封闭液为1%BSA+PBST,标准品用复溶液配制,抗体稀释液为1号,酶标二抗稀释液为3号,显色时间为15 min。本研究建立了一种灵敏、特异、操作简单、适于大批量样品筛选的SAL检测方法,对其他药物残留免疫学快速检测产品的研制及应用也有一定的借鉴意义。

By optimizing the ELISA working conditions, an indirect competitive ELISA method was established for detection of salbutamol （SAL）. The most suitable working condition of ELISA was obtained by phalanx titration method which is used to calculate the most optimal dilution of the coating antigen, antiserum, and HRP secondary antibody. Serial diluted SAL of the standard solution was set as the detection object and the standard curve of ELISA was plotted. The most optimal dilution of the coating antigen, antiserum, and HRP secondary antibody were 1：2000, 1：4000 and 1：6000, respectively. The standard curve of ELISA indicated that the linear determination was 0.5-20 ng/mL with 50% inhibitive concentration （IC50） of 2.33 ng/mL. The ELISA plates produced by Shenzhen Jincanhua Limited was chosen as the the best plate. And the optimization conditions were as follows： envelope liquid 0.05 moFL, pH 9.6 of carbonate buffer solution and blocking solution 1% BSA ＋ PBST. The standard substance was preparedwith combination solution Antibody diluent was of sample 1 and enzyme mark two anti diluents was of sample 3. coloration time was 15 minutes. This method for SAL detection was sensitive, specific, simple operation, suitable for screening large numbers of samples.