Starch gel electrophoresis of enzymes and polyacrylamide gel electrophoresis of muscle proteins were used to study genetic variation in 589 barramundi from three widely separated regions in northern Australia. Presumed genetic variation was observed at 16 loci (out of 46 screened). However, only 11 of these were polymorphic at the p0.99 level. The average heterozygosity in this species was 0.032. In nearly all cases, genotypic proportions agreed closely with Hardy-Weinberg expectations. Repeated sampling in each region revealed little or no change in allele frequencies over a period of several months. Similarly, comparisons of allele frequencies for fish from marine and from freshwater localities in the south-eastem Gulf of Carpentaria failed to reveal significant genetic differences between habitats. Between-region heterogeneity Χ2 tests indicated substantial genetic differentiation at 10 of the 11I polymorphic loci. These data indicate the existence of at least three distinct stocks or subpopulations of barramundi in Australia.