Bioassay of bile acids using an enzyme-linked DNA aptamer

• Published in: Analyst (London) Vol. 125; no. 8; p. 1371
• Main Authors: Kato, T, Yano, K, Ikebukuro, K, Karube, I
• Format: Journal Article
• Language: English
• Published: England 01-01-2000
• Subjects: Animals; Bile Acids and Salts - analysis; Biological Assay - methods; Cholic Acid - metabolism; DNA - metabolism; Ligands
• ISSN: 0003-2654
• DOI: 10.1039/b004580m

A new analytical method for the detection of bile acids has been developed by adopting an alkaline phosphatase-linked DNA oligomer that binds to bile acids. A 5'-biotin-labeled DNA oligomer with a 40-nucleotide length that is defined by the in vitro selection method was connected with alkaline phosphatase through an avidin-biotin linkage and applied to an enzyme immunoassay format. Sample solutions were incubated with small aliquots for a cholic acid-immobilized agarose matrix, on which the alkaline phosphatase-linked DNA oligomer had been bound prior to carrying out the assay. The amount of the alkaline phosphatase-linked DNA oligomer dissociated from the cholic acid-immobilized agarose matrix, which was detected using a fluorogenic substrate for alkaline phosphatase, indicated the amount of bile acids in the samples. The results suggest that the DNA aptamer directly linked with the reporter enzyme is applicable as a detector ligand for the immunoassay format. A linear calibration range was obtained for cholic acid between 0.1 to 5 mmol l-1 with a limit of detection of 10 mumol l-1. The %RSD was 7 at 5 mmol l-1 of cholic acid.