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金银花是常用大宗中药材之一,市场流通量大,但存在较多质量问题,建立一种金银花质量快速检测方法具有重要意义。木犀草苷是其质量控制指标之一,目前木犀草苷的含量检测主要以仪器检测为主,无法满足简单、快速、高通量的需求。酶联免疫检测法(ELISA)则具有灵敏度高、操作简单、设备仪器需求少、通量高等优点,是实现中药材有效成分快速检测的有效手段之一。该研究以木犀草素-7-O-β-D-葡萄糖醛酸苷为原料,通过活泼酯法,分别与牛血清白蛋白(BSA)及鸡卵清白蛋白(OVA)偶联制备免疫抗原及包被抗原;紫外分光光度法检测半抗原与载体蛋白的偶联比,分别为3.7∶1(LG-BSA)和1.0∶1(LG-OVA);免疫原(LG-BSA)免疫Bal b/c雌性小鼠,制备抗血清,ELISA检测抗血清效价并建立血清特异性标线。结果显示血清效价>2 000,以抗血清建立特异性标线,IC_(50)=298.7μg·L~(-1),标线范围18.4~4 852.4μg·L~(-1),表明所制备的完全抗原免疫原性良好。
Honeysuckle is one of the commonly used bulk Chinese herbal medicines, the market liquidity, but there are more quality problems, the establishment of a rapid detection of honeysuckle quality is of great significance. Luteolin is one of its quality control indicators. At present, the content of luteolin is mainly detected by instruments, which can not meet the demand of simple, rapid and high-throughput. Enzyme-linked immunosorbent assay (ELISA) has the advantages of high sensitivity, simple operation, less equipment requirements and high throughput, which is one of the effective means to achieve rapid detection of the active ingredients of Chinese herbal medicines. In this study, luteolin-7-O-β-D-glucuronide was used as the raw material to prepare immunological antigen by coupling with bovine serum albumin (BSA) and chicken ovalbumin (OVA) (LG-BSA) and 1.0: 1 (LG-OVA) respectively. Immunization with Balb / c Female mice were used to prepare antiserum. ELISA was used to detect antiserum titer and serum-specific labeling was established. The results showed that the serum titer was> 2,000, and the specific standard line was established by antiserum with IC 50 = 298.7 μg · L -1 and the standard range of 18.4-4.852.4 μg · L -1 The prepared complete antigen is immunogenic.