Abdul Waheed and Michael A. Moxley
The present invention generally relates to compositions and methods of detecting molecular interactions, such as protein interaction, in the fields of research, medical diagnostics, and DNA research. Various methods have been developed to study steady state levels, biosynthesis and turnover, mutation and stability, and effects of pharmacological compounds on expression. It is directed to compositions and methods for improving a chemiluminescent signal. In particular, a reaction buffer with an alkaline pH range of about 9 to 10 provides a maximal chemiluminescent intensity and longevity of luminol that is catalyzed by superoxide anions. The addition of carbonate to the reaction buffer provides an optimal signal to background ratio and stability of a chemiluminescent signal from luminol catalyzed by superoxide anions. The method includes preparing a buffer with an alkaline pH, combining the buffer with a working reagent to produce and detect a chemiluminescent signal. The working reagent includes luminol, a coumaric acid and a peroxide. The composition includes a buffer having a pH from about 9 to about 10, a stock reagent comprising luminol, a coumaric acid, a peroxide, and a second buffer having a pH of about 8.5.