A new strategy for the preparation of antibody against natural glycoside: With astragaloside IV as an example

Abstract

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"-CH₂OH 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 -NH₂ 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, IC₅₀, 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 (NaIO₄) 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 NaIO₄ oxidation. This new strategy provides new ideas for the design of haptens of other natural glycosides, as well as the preparation of their antibodies.

Keywords: Antibody; Artificial antigen; Astragaloside IV; Hapten; Natural glycoside.