Significance of Bilirubin as Anti-Oxidant in Gastrointestinal Surgery

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A new anti-bilirubin monoclinal antibody has been prepared and characterized. Using modified MBS (metamaleimidobenzoyl N-hydroxysuccinimide ester) method, haptenic unconjugated bilirubin-IXalpha (UCB) covalently coupled to bovine serum albumin (BSA) retaining its intramolecular hydrogen bonds and three-dimensional structure. After somatic cell fusion and screeing assay, we obtained anti-bilirubin monoclonal antibodies which reacted with UCB but did not react with bilirubin diglucuronide or monoglucuronide.To determine extremely low concentrations of bilirubin in human serum or urine, we utilized an antibody presently available (5M2) and another anti-bilirubin monoclinal antibody (24G7) which reacted with UCB as well as with conjugated bilirubin for enzyme-linked immunosorbent assay (ELISA). Bilirubin concentration in serm specimens obtained from many patients, determined using ELISA was compared with that determined by the conventional diazo method. When 5M2 was used in ELIZA, there was no discrepancy between bilirubin concentration determined by the two method in any sample. However, when 24G7 was used in ELISA, in the case of patients whose total urine volume was decreased, there was a significant discrepacy. This suggests the existence of a degraded bilirubin, substance "X", that is increased in the serum when any kind of stress occurs which will be excreted in the urine, In addition existence of substance "X" in the urine of healthy patients was observed.