STOPPED-FLOW FLUOROMETRY WITH AVAILABLE INSTRUMENTATION

Stopped-flow fluorescence experiments were conducted on a microscale in a spectrofluorometer by using an air-driven mixer attachment. All parts of the system employed, including the electronic components, are commercially available. There are many biochemical reactions of interest which take place in 0.05 to 10 seconds, that can be studied by fluorescence in this simple system. Examples shown are: (1) combination of carbonic anhydrase with a fluorescent sulfonamide; (2) reduction of α-ketoglutarate catalyzed by glutamate dehydrogenase at high concentration (1.5 mg/ml); and (3) formation of a ternary complex of lactate dehydrogenase. The mixing accessory is based on the design of R. Harvey and is compatible with a number of spectrometers. © 1969.