Fifty-seven mouse strains were examined for genetic variation at 21 esterase loci. Three new alleles were found: Es-6d in strain A/WySna, Es-lle in FTC/CpbU and Es-18c in two WLL/BrA sublines. At most loci there was a single allele found in over 80% of strains, with one or two rare alleles. However, the Es-1, 3, 10, 13, 25 and 27 loci were much more polymorphic. Although several loci were linked on chomosomes 3, 8 and 9, linkage disequilibrium was only found between Es-5 and Es-11 (chromosome 8) and Es-26 and Es-27 (chromosome 3). There was also significant disequilibrium between Es-1 and 3, Es-1 and 10, and Es-3 and 10, which are on different chromosomes, suggesting that the 57 strains are not a random sample of inbred mouse strains. Fifty-four strains were closely related, with the Es-7b, –17a, –18a, –23c set of alleles, which are typical of Mus musculus domesticus. The three exceptional strains were MOL3 (Mus musculus molossinus), WLL/BrA (English–Norwegian origin) and TA2 (Chinese origin). There were 10 groups of strains which were identical at all loci. Sublines of the same strain were usually identical. Sometimes more distantly related strains, such as CBA/Bi, C3H/He, SM and DBA/Li, were identical, and in a few cases strains with no known common ancestry such as C58 and MAS were identical. Attempts to discriminate between a subset of 22 American and 15 European strains were unsuccessful, suggesting that the European strains add only in a quantitative manner to the gene pool of ‘laboratory mice’, whereas wild-derived strains such as MOL3 are genetically quite distinct from other laboratory mice.