Bilirubin-ProteinInteractionsMonitoredby Difference Spectroscopy

Difference spectroscopy is used to monitor bilirubin-protein interactions, to assess the residual binding capacity of proteins for bilirubin. A change in the difference spectra monitored at 482 nm is directly proportional to bound bi-lirubin up to a molar ratio of bilirubin to albumin of ap-proximately 1; increasing bilirubin beyond the 1:1 molar ratio does not further changethe difference spectra. After excess free bilirubin is added, the change in the difference spectrum is proportional to the residual binding capacity of the serum for bilirubin. The risk of kernicterus among neonates may be assessed by monitoring the residual bi-lirubin binding capacity of serum. This report summarizes our research effort leading to an assay method which re-quires only 40 il of serum and can be completed in less than 10 mm