Development of Sensitive Monoclonal Antibody-based Enzyme-linked Immunosorbent Assay for Renal Toxicity Ingredient Aristolochic acid A in Chinese Herb Medicine
Aristolochic acid A-bovine serum albumin (BSA) conjugate and aristolochic acid A-ovalbumin (OVA) conjugate,were produced as the immunogen and coating antigen,respectively. A monoclonal antibody (mAb),designated as 1A11,was produced with aristolochic acid A-BSA. The titer was 2×104. The mAb was IgG1 isotype with κ light chains. A sensitive monoclonal antibody-based indirected enzyme-linked immunosorbent assay (icELISA) was established with the mAb. The cross-reactivity (CR) of mAb 1A11 with aristolochic acid B,C and D,an analog of aristolochic acid A,was 2.8%,3.5% and 31.2%,respectively. IC50 and the working range of the icELISA for aristolochic acid A were 1.9 and 0.5-7.5 μg/L,respectively. The average recoveries of aristolochic acid A fortified in caulis clematidis armandii were 86%-97% with RSD of 5.2%-11.1%. Analysis of aristolochic acid A in Chinese herb medicines and Chinese proprietary medicines with icELISA showed that the examined samples of manshurian dutchmanspipe stem,herba aristolochiae,fructus aristolochiae,aristolochia fangchi,and dutchmanspipe root were positive,the caulis clematidis armandii and the other Chinese proprietary medicines examined samples were negative and the results were confirmed by HPLC analysis. This ELISA is suitable for the rapid detection of aristolochic acid A in Chinese Herb medicine.