A new strategy for the hapten design of natural glycoside and application for the preparation of antibody is reported in this work. With astragaloside IV (AGS-IV) as an example, C6"-CHOH on a glucosyl group was selectively oxidized by 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) oxidation to C6"-COOH, which was subsequently condensed with -NH on bovine serum albumin to get artificial antigen. Then, the successful preparation of artificial antigen was verified by TCL, SDS-PAGE, UV, and MALDI-TOF-MS. Finally, rabbits were immunized with artificial antigen to obtain an antibody against AGS-IV. After tests of the titer, IC, and cross-reactivity, the results showed that the antibody prepared by TEMPO oxidation in this work had higher specificity than that the antibody prepared by conventional sodium periodate (NaIO) oxidation. The hapten, as a carboxylic acid derivative of AGS-IV, has better water solubility than AGS IV, which is more suitable for the synthesis of the hapten-carrier protein conjugate in aqueous phase, achieving another virtue of TEMPO oxidation over NaIO oxidation. This new strategy provides new ideas for the design of haptens of other natural glycosides, as well as the preparation of their antibodies.
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