Ultramicro Method for Serum Bilirubin by Diazo Blue Reaction

Abstract An ultramicromethod is presented for the determination of serum bilirubin by the (acid) diazo blue reaction, using heel puncture or fingertip blood. Diazotization is accomplished in a strongly acid medium with the resulting blue azobilirubin having a maximum absorption at 580 mµ. The method is extremely reproducible and sensitive, being carried out in final serum dilutions of 1:100 to 1:40. The method is described for use with the Coleman Junior spectrophotometer and can be adapted to other instruments. The diazo blue micromethod compares very favorably with the Malloy and Evelyn method which was done in parallel. The results of this study also suggest that the diazo blue micromethod can be used on venous serum or serum obtained from fingertip puncture without having markedly altered results.