Reserve bilirubin binding capacity determined by difference spectroscopy. I. Modifications and performance of the difference spectroscopy assay.

The difference spectroscopy assay for reserve bilirubin binding capacity, modified to improve its performance, was used to study solutions of human serum albumin and bilirubin. The results indicate that the assay is performed simply and rapidly; that it uses very small sample volumes, making it suitable for use in premature human neonates; that it measures reserve binding capacity and not free bilirubin resulting from exceeding binding capacity; that it measures binding at two sites, which can be discriminated by blocking the low-affinity site with concentrated salicylate; and that it can be performed at varying pH depending upon the experimental need.