This study was conducted to develop a proficiency test (PT) specimen for the enumeration of Escherichia coli, a sanitary indicative bacterium, in molluscan bivalve shellfish. The common mussel Mytilus edulis was chosen as a matrix for the PT specimen. Mussels were artificially bioaccumulated for approximately 3 h with E. coli culture. After determining the homogeneity of samples, samples were distributed to 17 participants involved in the proficiency testing program. The enumeration of E. coli was performed by the most probable number method of the American Public Health Association. Statistical evaluations of the results obtained from inter- and intra-laboratory variation indicated no significant differences in the accuracy of these techniques between participants, indicating z-scores of ${leq}{pm}2$ and suggesting that preparation of the PT specimen for enumerating E coli in shellfish was successful.