摘要
目的合成绿原酸（chlorogenic acid，CHA）人工抗原，制备CHA多克隆抗体，为制备过敏成分分析的免疫芯片奠定基础。方法采用碳二二亚胺法分别合成CHA免疫抗原（CHA-牛血清白蛋白）和包被抗原（CHA-卯清蛋白）；UV泫和TLC法鉴定人工抗原偶联是否成功；间接ELISA方法检测免疫小鼠血清中抗体效价；间接竞争ELISA方法检测抗体特异性。结果经UV法和TLC法检测，CHA与BSA偶联成功。血清抗体效价为1：128000，CHA-PAbs的质晕浓度与抑制率的线性万程为Y=一0．101nX＋1．158，线性检测范围为15．6～250ng／mL，R^2=0．995，并且和甘草酸、黄芩苷等与其常配伍应用的中药化合物均无交义反应。结论成功制备了灵敏度高、特应性好的CHA多克隆抗体，为CHA的微量检测和过敏性研究提供了实验依据。
Objective To synthesize artificial chlorogenic acid （CHA） antigen, to prepare CHA polyclonal antibody, and to lay the foundation for the preparation of immune chip for allergic component analysis. Methods CHA-bovine serum albumin （CHA-BSA） and CHA-ovalbumin （CHA-OVA） were synthesized by carbodiimide method. The characterization of the synthesis was examined by UV spectrometry and TLC method. The titer and specificity of the antibody in serum of immunised mice were detected by indirect enzyme-linked immunosorbent assay （I-ELISA） and indirect competitive enzyme-linked immunosorbent assay （I-CELISA）, respectively. Results According to the UV and TLC determination, the CHA was successfully conjugated with BSA. The serum antibody titer was 1 ： 128 000 with the linear range of 15.6-50 ng/mL, R^2 - 0.995, and Y=-0. 10 In）（＋ 1. 158. No cross-reaction was detected with glycyrrhizic acid, parietic acid and so on, which were commonly used in the combination with CHA. Conclusion The preparation of multi-clonal antibody with good sensitivity and specificity against CHA is successful, which provides a foundation for the trace detection and allergy research of CHA.
出处
《中草药》
CAS
CSCD
北大核心
2014年第4期480-484,共5页
Chinese Traditional and Herbal Drugs
基金
国家自然科学基金资助项目(81274043
81373542)
"重大新药创制"国家专项课题(2010ZX09502-002)
关键词
绿原酸
人工抗原
多克隆抗体
碳二亚胺法
免疫分析
chlorogenic acid
artificial antigen
polyclonal antibody
carbodiimide method
immune analysis
