Immobilized bacterial luciferase for microscale analysis of creatine kinase activity

The commercially available bacterial luciferase: oxidoreductase system obtained from Vibrio fischerii has been immobilized in a bovine serum albumin (BSA) gel. The gel was cut in the shape of a disc and held to the bottom of a reaction cell, gel upwards. The immobilized enzyme gels are stable, reusable and easily cleaned of spent reagents. NADH and NADPH have been assayed down to nanomolar concentrations, although with an error of ± 15%. The system has been coupled to an NADPH-producing commercial assay for creatine kinase (ATP: creatine N-phosphotransferase, EC 2.7.3.2) activity. The kinetic assay gives a linear reaction rate vs. creatine kinase activity plot in the clinically important range.