Nitroimidazoles (NDZs) Hapten Design and Synthesis
Based on the mature experience in the development of small molecule drugs and antibodies, Creative Biolabs can provide customers with hapten design and modification services for antibiotics, food additives, heavy metals, toxins, etc. to help customers develop corresponding antibodies. The antibodies we design and develop have good quality and can be perfectly applied to downstream related detection.
Nitroimidazoles (NDZs) are classified as illegal drugs in many countries because of their mutagenicity and potential carcinogenicity. In the European Union, Ronidazole (RNZ), Dimetridazole (DMZ), and Metronidazole (MNZ) were banned from use in food animals in 1993, 1995, and 1998, respectively, and the FDA in 2002 announced 11 drugs banned from use in imported foods of animal origin, including DMZ and other nitroimidazole drugs. Therefore, there is an urgent need to establish a detection method that is low in cost, quick and easy to operate, and can guarantee accurate and reliable results. One such method is immunological tests based on specific binding of antigens and antibodies.
NDZs are a class of drugs with a 5-nitroimidazole ring structure. The main varieties include metronidazole (MNZ), dimethyl metronidazole (DMZ), ipronidazole (IPZ), secnidazole (SCZ), ornidazole (ONZ), tinidazole (TNZ) and ronidazole (RNZ) and the like. NDZs have antiprotozoal and antibacterial activity. After entering susceptible microbial cells, in an anaerobic or hypoxic environment and low redox potential, the nitro group of NDZs is easily reduced by electron transfer proteins to cytotoxic amino groups, which inhibits cell DNA synthesis, destroy the double-helix structure of DNA or block its transcription and replication, thereby causing the cell to die, and play its role in quickly killing anaerobic bacteria and effectively controlling infection. Huet et al. have established ELISA methods that can detect RNZ, DMZ, HMMNI, MNZ, and IPZ in chicken and eggs. The detection limits of DMZ in eggs and chicken are 1 μg/kg and 2 μg/kg, respectively. The detection limit of MNZ is less than 10 μg/kg, RNZ and HMMNI are less than 20 ug/kg, while IPZ is less than 40 ug/kg. Creative Biolabs can provide class-specific and compound-specific NDZs hapten design and modification services to meet customers' development needs for broad-specific or certain NDZs antibodies.
Fig.1 Chemical structures of typical NDZs and metabolites. (Chuanlai Xu, 2019)
Successful class-specific hapten design can develop broad-specific NDZs antibodies, so several NDZs residues can be detected in a single sample. Therefore, by designing and optimizing the mother ring structure of NDZs, and selecting appropriate carriers and ligation methods, a complete antigen with both immunogenicity and reactivity can be prepared, and then class-specific antibodies can be produced. 1D5 is such a monoclonal antibody with IC50 of 4.79 µg/L, 0.47 µg/L, 5.97 µg/L, 23.48 µg/L, and 15.03 µg for dimetridazole, ipronidazole, ronidazole, hydroxydimetridazole, and hydroxyipronidazole, respectively. The detection limit of NDZs established by the 1D5-based ELISA method was 4.2 to 50.3 µg/kg in feed matrices and 0.11 to 4.11 µg/kg in edible animal tissues. In addition, the results show that the ELISA method established with this antibody has a good correlation with the conventional HPLC-MS method (r2 = 0.9905).
Although the use of class-specific antibodies to detect multiple NDZs in a single sample can avoid the trouble of using mixed antibodies, sometimes broad-spectrum antibodies can cross-react with multiple antigens/drugs. So, it is difficult to guarantee its complete specificity for a certain test molecule, and this problem is particularly intractable when a quantitative analysis of a certain drug or residue is required. Therefore, it is also necessary to design compound-specific haptens to develop antibodies with high specificity for a certain compound.
Creative Biolabs has rich experience in hapten design and modification. The antibodies we design and develop can fully meet customers' testing needs and have been well received and praised by customers worldwide. If additional help is needed, please directly contact us and consult our technical supports for more details.
- Chuanlai Xu.; et al. Food Immunoassay. Chapter 6, Antibiotics Immunoassay in Food, Page 224.