狂犬病病毒抗体胶体金检测试纸的制备

Preparation and application of a colloidal gold strip to detect the rabies antibody

摘要:

通过胶体金免疫层析技术建立一种特异、便捷、快速的狂犬病病毒抗体检测方法，对犬等动物免疫狂犬病疫苗后的抗体水平监测提供参考。用醋酸锌沉淀法沉淀狂犬病病毒CVS11，Sepharose 4FF进行层析纯化。用柠檬酸三钠还原法制备的胶体金，标记纯化的狂犬病病毒，喷于试纸的结合释放垫 (金标垫)，将SPA (葡萄球菌表面A蛋白) 和纯化的兔抗狂犬病病毒IgG分别喷于试纸的T (检测线) 处和C (对照线) 处，组装试纸条。用制备的试纸条对261份犬血清进行检测，与快速荧光灶抑制试验 (RFFIT) 检测的结果一致；对已知效价的犬狂犬病毒中和抗体 (VNA) 大于0.5 IU，结果为阳性；对狂犬病病毒中和抗体 (VNA) 小于0.5 IU/mL的血清，结果为阴性。制备的狂犬病病毒抗体胶体金检测试纸检测犬血清抗体，具有特异、便捷、快速的特点，能够检测出狂犬病病毒中和抗体大于0.5 IU的血清，适用于临床犬血清抗体水平监测，具有良好的应用前景。

Abstract:

To develop a specific, rapid, and convenient immunochromatography assay (ICA) to detect the rabies antibody in clinical sample from immuned dogs by rabies vaccine. Colloidal gold particles labeled with purified rabies virus (CVS11) were used as the detector reagent. The staphylococcal protein A (SPA) and pured rabbit anti-rabies virus IgG were blotted on the test and control regions of nitrocellulose membrane. Then the strip was assembled with sample pad, absorbing pad, and dorsal shield. The assay samples (261 dog’s serum) were collected from Wildlife Rabies Disease Diagnostic Laboratories of Ministry of Agriculture in China, Institute of Military Veterinary, Academy of Military Medical Sciences and other six provinces, including rabies virus positive and negative serum. The performance of the strip was compared to fluorescent antibody virus neutralization test. The neutralizing antibody titer could be detected above 0.5 IU. The strip did not change of performance when stored at room temperature for 12 months. It may offer reference of neutralizing antibody titer level after dogs immuned rabies vaccine and determin whether the dogs need to be immuned again.