Anti-bilirubin monoclonal antibody II. Enzyme-linked immunosorbent assay for bilirubin fractions by combination of two monoclonal antibodies

Yasuharu Izumi, Masahiko Yamazaki, Shuji Shimizu, Kuniko Shimizu, Tokio Yamaguchi, Hiroshi Nakajima

Research output: Contribution to journalArticle

41 Citations (Scopus)

Abstract

Among 16 monoclonal antibodies raised against covalently coupled bilirubin-bovine serum albumin, we selected two antibodies: one (designated 24G7) reacted with unconjugated and conjugated bilirubin and the other (designated 25H17) reacted only with unconjugated bilirubin. Combination of these two antibodies enabled us to determine extremely low concentrations of unconjugated and conjugated bilirubin independently by enzyme-linked immunosorbent assay (ELISA). In the assay, samples were incubated with each anti-bilirubin IgG, and then free remaining IgG was allowed to bind to the immunotiter plates coated with bilirubin-bovine serum albumin. The bound fraction of the IgG was visualized with horseradish peroxidase-conjugated rabbit anti-mouse IgG and substrate. Bilirubin concentration was determined from the absorbance at 425 nm. In this system, we could measure 10-7-10-5 M unconjugated and conjugated bilirubin in samples, which is 100-fold more sensitive than Michaëlsson's diazocoupling method. The assay results gave a good correlation coefficient (0.86) compared with those determined with high performance liquid chromatography.

Original languageEnglish
Pages (from-to)261-266
Number of pages6
JournalBBA - General Subjects
Volume967
Issue number2
DOIs
Publication statusPublished - Nov 17 1988

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology

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