In several laboratory and production species, the establishment of a proteome reference map of a specific tissue has been accomplished. The rabbit is widely used as both a production and experimental animal. A lot of physiology research involving the gastrocnemius muscle of rabbit is described, although no reference proteome map is available. In this work, the first reference map of the rabbit's gastrocnemius muscle using 2D gel electrophoresis and the identification of proteins through peptide mass fingerprinting (PMF) was established. A total of 45 proteins were localized and identified with three major roles: cell structure and contractile apparatus: metabolic and cell defense proteins. A reference map of major proteins expressed is described enabling possible comparisons with other physiological studies.