In Drosophila, the concentration and phosphorylation levels of several important circadian proteins (e.g., PERIOD, TIMELESS) oscillate on a 24-h basis. A simple and rapid method for extracting proteins from fly heads is presented here. The extracts can immediately be loaded onto an sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel to assay the effects of mutations, genetic manipulations, or environmental conditions on the oscillations of circadian proteins by Western blotting. They can also be used for immunoprecipitation experiments.