Bupleurum Saponins A Artificial Antigen Synthesis And Preparation Of Monoclonal Antibodies

Saikosapoins a (SSa) is a natural compound mainly isolated from Bupleurum Chinese DC or Bupleurum scorzonerifolium Willd. As a traditional Chinese medicine in china, the quantification of SSa has great significance in the study of Traditional Chinese Medicine (TCM). The concentration of saikosaponins(SS) is suggested to be the important piece of pharmacological study. The complicated pretreatment remains indispensable in traditional analytical methods. Comparing with these methods, ELISA is a more simpler and more environmentally determination method. It plays a very important role in pharmacological study of Traditional Chinese MedicineMethod:Artificial antigents for immunization (SSa-BSA) and coating (SSa-PLL) were synthesized through sodium periodate oxidization. The reaction product is identified by UV and TLC after dialyzed for2days.We take Balb/C male mice as immune animals. After the fifth immunization, we measured the blood serum of the Balb/C mice. The titers were determined by indirect ELISA, and the specificity was determined by the complete indirect ELISA.The spleen cells from the well immunized animals were fused with myeloma cells. After three times of limiting dilution, we obtained hybridoma lines which can secrete Mab against SSa. These cell lines were injected into mice’s abdominal cavity to induce MabsUsing the method of indirect competitive elisa (IC-ELISA),we detected its Specificity, sensitivity and cross-reactivity with other compounds.Result: The results showed that SSa and carrier proteins were condensated successfully. The title of anti-serum is above1:80000. The detection results showed that the specificity of McAb was high and the cross-reactivity with other compounds was low. The standard curve equation was y=8.2347x-9.0765, R2=0.9962. The Linear Range was at1-500μg/ml.