A new method for total, free, and conjugated serum bilirubin.

Abstract

Methods for the measurement of conju­ gated and free serum bilirubin have attracted the attention of investigators for almost half a century. Such estimations would more ac­ curately reflect the true content of these two major components of bilirubin than do re­ sults now available with the use of diazo methods. The accurate estimation of these two bili­ rubin fractions in serum would aid in more accurate assessment of diseases associated with abnormal amounts of these fractions. It might also be useful in the detection of hepatic functional impairment in the pres­ ence of a normal total serum bilirubin. 5 The diazo system 19 ' 23 - 24 for estimating these two bilirubin fractions has been criti­ cized for its inability to differentiate direct from indirect serum bilirubin. Investigators 6,10, 16 report that evidence does not support the contention that the 1min. fraction of the diazo reaction is a measure of a distinct type of bilirubin. The evidence suggests that the rate of diazotization is determined by the concentration of the bilirubin and by chemical or physical factors in serum, and not by the presence of two types of bilirubin with different reaction rates. Hemobilirubin is present in normal blood serum and is increased in cases of hemolytic jaundice. It gives the indirect van den Bergh reaction and is readily extracted by chloroform. 11 The cholebilirubin present in bile and in serum in cases of hepatic or ob­ structive jaundice gives the direct van den Bergh reaction and is not extracted by chloroform. The differences in the solubility in chloroform between the hemobilirubin (free) and cholebilirubin (conjugated) permit separation of these two types of pigment in the serum. 11