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@C82(OH)x 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.
Keywords: Enzyme immunoassay; Immunogen; Metallofullerene; Nanomedicine; Polyclonal antibody.