LSP-1 beta S is present in Michigan and Massachusetts Drosophila melanogaster natural populations. Its frequency, 10%, is significantly higher in an East Jordan, Mich. (latitude, 45.10 degrees N), population than in East Lansing, Mich. (latitude 42.44 degrees N), or Hadley, Mass. (latitude, 42.21 degrees N), populations, where it averages 3% at each location. The average frequency of LSP-2S is more comparable, 6, 5, and 7% at East Jordan, East Lansing, and Hadley, respectively. LSP-1 gamma F variants are also present. A total of 342 single third-instar larvae was scored for LSP-1 autosomal variants, and 323 for LSP-2 variants. Each larva represented a newly established isofemale line from collections at East Jordan in 1981 and 1983, East Lansing in 1982, and Hadley in 1981, 1982, and 1983. Within localities, frequencies of hemolymph protein variants did not differ significantly between years. Proteins 9, 10, 11, and 15 correspond to the LSP-1 gamma, beta, and alpha triplet and LSP-2 polypeptide in D. melanogaster. Our results together with those of Singh and Coulthart [(1982). Genetics 102:437] indicate that D. melanogaster populations in north temperate climates maintain considerable genetic heterogeneity for the larval hemolymph proteins.