Abstract
Herein is reported an immunochemical approach to determine hyodeoxycholic acid using hybridoma-secreting monoclonal antibodies. The hyodeoxycholic acid-specific antibody was produced by fusing splenocytes immunized with a hyodeoxycholic acid-bovine serum albumin conjugate with a hypoxanthine–aminopterin–thymidine-sensitive mouse myeloma cell line (SP2/0). The antibody was highly specific for hyodeoxycholic acid, with less than 0.05 percent cross-reactivity to over fifty structurally related compounds. The antihyodeoxycholic acid monoclonal antibody was then used to develop a rapid, specific, and sensitive indirect competitive enzyme-linked immunosorbent assay (icELISA) for the determination of hyodeoxycholic acid in pharmaceutical compounds. The linear dynamic range was from 0.48 to 62.5 nanograms per milliliter with an IC50 value of 8 nanograms per milliliter. The icELISA results correlated well with a conventional high-performance liquid chromatography method for the determination of hyodeoxycholic acid (R2 = 0.9982). This study shows that the icELISA method was successfully applied to the quantification of hyodeoxycholic acid in pharmaceutical products.
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