CAJ | 학술논문

目的建立特异、敏感的检测大肠杆菌菌体蛋白的夹心ELISA试剂盒.方法采用大肠杆菌菌体蛋白免疫家兔,制备得到高效价抗菌体蛋白抗血清.将经饱和硫酸铵盐析、阴离子交换层析和亲和层析纯化的兔抗大肠杆菌菌体蛋白特异性多克隆抗体作为包被抗体和检测抗体,用生物素标记检测抗体,并加入辣根过氧化物酶标记的亲和素.结果建立的大肠杆菌菌体蛋白夹心ELISA检测方法的敏感性为0.32 μg/L,检测范围为1～100 μg/L,与国际同类商品化试剂盒相当.此法具有良好的稳定性,其批内变异系数小于7.7%,批间变异系数小于6.2%.结论建立了特异、敏感、稳定的检测大肠杆菌菌体蛋白的夹心ELISA试剂盒,为生物制品中残留大肠杆菌菌体蛋白的质量控制提供了一种重要的检测方法.
목적 건립특이、민감적검측대장간균균체단백적협심ELISA시제합.방법 채용대장간균균체단백면역가토,제비득도고효개항균체단백항혈청.장경포화류산안염석、음리자교환층석화친화층석순화적토항대장간균균체단백특이성다극륭항체작위포피항체화검측항체,용생물소표기검측항체,병가입랄근과양화물매표기적친화소.결과 건립적대장간균균체단백협심ELISA검측방법적민감성위0.32 μg/L,검측범위위1～100 μg/L,여국제동류상품화시제합상당.차법구유량호적은정성,기비내변이계수소우7.7%,비간변이계수소우6.2%.결론 건립료특이、민감、은정적검측대장간균균체단백적협심ELISA시제합,위생물제품중잔류대장간균균체단백적질량공제제공료일충중요적검측방법.
Objective To establish a highly specific and sensitive sandwich ELISA kit for detection of E. coli cell proteins. Methods E. coli cell proteins were used for immunization of rabbits and high titer antisera were obtained. The polyclonal antibodies(PAbs)against E. coli cell proteins were purified by saturated ammonium sulfate salting-out, anion exchange chromatography and affinity chromatography. The purified PAbs were used as coating antibodies, and biotin-labeled PAbs plus horseradish peroxidase-conjugated avidin were used as detecting antibodies. Results The sensitivity and detection range of the sandwich ELISA system for detection of E. coli cell proteins were 0.32 μg/L and 1-100 μg/L, respectively, and equal to the similar commercial kits in the world. This ELISA kit had good stability, and its intra- and inter-assay variation coefficients were less than 7.7% and less than 6. 2%, respectively. Conclusions The sandwich ELISA kit with high specificity, sensitivity and stability is established successfully and provides an important method for quality control of residual E. coli cell proteins in biological products.