title: An enzymatic cycling method for the determination of serum total bile acids with recombinant 3alpha-hydroxysteroid dehydrogenase.
abstract: A highly sensitive enzymatic cycling method was developed for the serum total bile acids assay. We constructed a prokaryotic expression system to prepare the recombinant 3alpha-hydroxysteroid dehydrogenase in place of the natural enzyme and for the first time used it in the total bile acids assay. The production rate of thio-NADH correlated with the bile acids concentration and was measured by the change of absorbance at 405/660 nm. The enzymatic cycling method could detect 0.22 micromol/L total bile acids in serum. Within-run and between-run imprecisions were 1.2-3.7% and 2.3-4.8%, respectively. The calibration curve for total bile acids in serum was linear between 0.5 and 180 micromol/L. This method was free from interference by bilirubin, hemoglobin, ascorbate, and lactate dehydrogenase. In conclusion, serum total bile acids could be measured by the enzymatic cycling method with recombinant 3alpha-hydroxysteroid dehydrogenase as the tool enzyme.
authors.affiliation: Department of Clinical Laboratory, Peking University First Hospital, Beijing, China.
authors.forename: Guo-Hua
authors.initials: GH
authors.lastname: Zhang
authors.forename: Ai-Ri
authors.initials: AR
authors.lastname: Cong
authors.forename: Guo-Bing
authors.initials: GB
authors.lastname: Xu
authors.forename: Chuan-Bao
authors.initials: CB
authors.lastname: Li
authors.forename: Rui-Feng
authors.initials: RF
authors.lastname: Yang
authors.forename: Tie-An
authors.initials: TA
authors.lastname: Xia
mesh_terms: D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific) / Q000737:chemistry* / Q000235:genetics; D000818:Animals; D001647:Bile Acids and Salts / Q000097:blood*; D001774:Blood Chemical Analysis / Q000379:methods*; D020604:Comamonas testosteroni / Q000201:enzymology* / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D011994:Recombinant Proteins / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D001647:Bile Acids and Salts; D011994:Recombinant Proteins; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific)