Immobilization of luciferase from a firefly lantern extract on glass strips as an alternative strategy for luminescent detection of ATP

The bioluminescent reaction catalysed by firefly luciferase has become wide ly established as an outstanding analytical system for assay of ATP. When u sed in solution, luciferase is unstable and cannot be re-used, a problem th at can be partially circumvented by immobilizing the enzyme on solid substr ates. Transparent glass is especially advantageous over alternative immobil izing matrices, since it allows most of the emitted photons to be detected. We report a new method for luciferase immobilization on glass which does n ot require prior silanization and glutaraldehyde activation, thus saving pr eparation time and minimizing enzyme inactivation. Our method is based on t he co-immobilization by adsorption of luciferase (from a firefly lantern ex tract) and poly-L-lysine (PL) on non-porous glass strips. Luciferase immobi lized in this way exhibits minimal variations in intersample activity, high sensitivity for ATP detection (linear luminescence responses down to 50 nm ol/L) and good stability (full activity for at least 60 days when stored at -80 degrees C). PL-mediated immobilization of luciferase on glass strips p rovides an attractive strategy for the design of specific ATP biosensors, w ith potential in industry, environmental screening, medicine and biological research. (C) 1998 John Wiley & Sons, Ltd.