Guo Xihong, Yang Shangyuan, Huang Huan, Cui Rongli, Li Cheng, Yao Huanli, Liu Bing, Zhang Lele, Xu Binggang, Dong Jinquan, Sun Baoyun
CAS Key Laboratory for Biomedical Effects of Nanomaterials and Nanosafety, Institute of High Energy Physics, Chinese Academy of Sciences, Beijing, 100049, China.
Anal Bioanal Chem. 2017 Nov;409(28):6575-6581. doi: 10.1007/s00216-017-0606-1. Epub 2017 Sep 22.
With their intriguing structures and properties, metallofullerenols have attracted considerable attention in biological and medical applications. Due to the increasing biomedical interest, effective detection methods are important to monitor and control metallofullerenols. However, the detection of metallofullerenols becomes very difficult after polyhydroxylated modification due to the lack of detectable features. Antibody-based immunoassay methods have been important tools for detection and will better meet the needs of analysis of metallofullerenols. Thus, the antigenicity of metallofullerenol has been studied for the first time. In this study, no immune response was detected when metallofullerenol Gd@C(OH) was used as immunogen. However, the polyclonal antibody against metallofullerenol was produced using metallofullerenol-KLH (keyhole limpet hemocyanin) as immunogen, indicating that metallofullerenol can act as hapten. The specificity of the obtained antibody was investigated. It has been found that the hydroxyl groups on the surface of the carbon cage, the encapsulated metal, and the size and shape of the carbon cage did not affect the recognition specificity of the antibody. Based on the obtained antibody, an indirect competitive enzyme immunoassay was developed for the determination of metallofullerenol with detection limits of 18 ng/mL in PBS. This enzyme immunoassay method was successfully used to detect metallofullerenol in serum. This work can provide an innovative way to determine metallofullerenols. Graphical abstract The polyclonal antibody against metallofullerenol was produced using metallofullerenol-KLH (keyhole limpet hemocyanin) as immunogen. Based on the obtained antibody, a competitive enzyme immunoassay was developed for the determination of metallofullerenol.
金属富勒醇因其引人入胜的结构和性质，在生物和医学应用中备受关注。由于生物医学领域的兴趣日益浓厚，有效的检测方法对于监测和控制金属富勒醇至关重要。然而，由于缺乏可检测特征，多羟基化修饰后的金属富勒醇检测变得非常困难。基于抗体的免疫分析方法一直是检测的重要工具，并且将更好地满足金属富勒醇分析的需求。因此，首次对金属富勒醇的抗原性进行了研究。在本研究中，当使用金属富勒醇Gd@C(OH)作为免疫原时，未检测到免疫反应。然而，以金属富勒醇-钥孔血蓝蛋白（KLH）作为免疫原产生了抗金属富勒醇的多克隆抗体，这表明金属富勒醇可以作为半抗原。对获得的抗体的特异性进行了研究。已经发现碳笼表面的羟基、包封的金属以及碳笼的大小和形状并不影响抗体的识别特异性。基于获得的抗体，开发了一种间接竞争酶免疫分析法用于测定金属富勒醇，在磷酸盐缓冲盐溶液（PBS）中的检测限为18 ng/mL。该酶免疫分析方法成功用于检测血清中的金属富勒醇。这项工作可为测定金属富勒醇提供一种创新方法。图形摘要：以金属富勒醇-钥孔血蓝蛋白（KLH）作为免疫原产生了抗金属富勒醇的多克隆抗体。基于获得的抗体，开发了一种竞争酶免疫分析法用于测定金属富勒醇。
