We have developed a rapid, automated assay for glycogen phosphorylase that measures the degradative reaction. The substrate contains higher concentrations of phosphate and AMP than other methods, and the enzyme is preincubated with the substrate before adding phosphate to start the reaction. These modifications improve the activity and linearity of the assay. The new assay compares favourably with a standard phosphorylase assay in the synthetic direction and is linear to 500 U/L. We have used this assay to measure phosphorylase activity in human tissue samples and muscle biopsy specimens.