绿磺隆克百威三唑磷多抗体免疫亲和色谱技术研究

采用碳酰二咪唑(CDI)将Sepharose CL-4B活化并分别与纯化的抗绿磺隆、抗克百威、抗三唑磷的多克隆抗体共价偶联,合成相应的免疫亲和吸附剂,并制备了对绿磺隆、克百威和三唑磷具有特异性亲和力的多抗体免疫亲和色谱(MIAC)柱.对MIAC的条件进行优化,选择0.02 mol·L~(-1) pH7.2磷酸盐缓冲液作吸附与平衡介质,80%(体积分数)甲醇水溶液作洗脱剂.结果表明,在上述试验条件下,MIAC柱对绿磺隆、克百威和I唑磷的动态柱容量分别达1.81、2.29和1.89μg·mL~(-1)床体积.用MIAC柱对添加有绿磺隆、克百威、三唑磷的河水与土壤提取液进行分离富集,洗脱液分别采用包被抗体直接竞争酶联免疫吸附(ELISA)法和高效液相色谱(HPLC)法测定,重复5次,平均回收率是89.98%～106.2%,相对标准偏差(RSD)为3.21%～14.81%,ELISA法和HPLC法的测定结果基本一致.成功建立了绿磺隆、克百威和三唑磷的多抗体免疫亲和色谱分析技术并用于河水和土壤中绿磺隆、克百威、三唑磷残留的测定.

Studies on Multi-class Immunoaffinity Chromatography(MIAC)for Chlorsulfuron, Carbofuran and Triaophos

The purified anti-chlorsulfuron antibody, anti- carbofuran antibody and anti-triazophos antibody were conjugated to the CDI-acti-vated sepharose CL-4B respectively to synthesize the immunosorbent. The mixed immunosorbent of chlorsulfuron, carbofuran and triaophos were used to prepare multi-class immunoaffinity chromatography( MIAC) column which are specific to chlorsulfuron, carbofuran and triaophos. The conditions of MIAC were optimized, 0.02 mol·L~(-1) pH 7.2 phosphate buffer was used as equilibrium and absorbent medium, and 80% methanol-water( V/V ) as eluent. Under the experimental conditions, the dynamic column capacity of chlorsulfuron, carbofuran and triaophos were up to 1.81, 2.29 and 1.89 μg·mL~(-1) bed volume respectively. The spiked river water and the extracts of spiked soil were clean up and en-riched by MIAC,the target compounds in eluates were determined by enzyme linked immunosorbent assay(ELISA) and verified by high per-formance liquid chromatography(HPLC). The average recoveries were 89.98%～106.2% and the RSD were 3.21%～14.81%(n=5). The results from ELISA were well corresponded with those from HPLC. MIAC technology was developed successfully for analysis of chlorsulfuron, carbo-furan and triaophos residues in water river and soil.