Allelic variation at fourteen loci coding for ten enzymes was studied in seventeen populations of Hyla regilla and in H. eximia, H. wrightorum, H. cadaverina, and H. euphorbiacea. Heterozygote deficiencies were found at several loci in several populations of H. regilla and possible reasons are suggested. The proportion of polymorphic loci and average individual heterozygosity were typical of other vertebrates with some exceptions: (1) the Lee's Camp, Oregon population; (2) the Santa Cruz Island population; (3) the three San Diego Co. populations; (4) the Sierra San Pedro Martyr population; and (5) the three populations in southern Baja California. Measures of genetic similarity (S) and genetic distance (D), as well as patterns of variation in allele frequencies at PGI and IDH-2, suggested three groups within Hyla regilla. These were: (1) the four Oregon populations; (2) the three central California populations; and (3) the ten southern California and Baja populations. The proportion of major alleles shared indicated Hyla eximia and H. wrightorum are very closely related and that H. wrightorum is not a subspecies of H. regilla; H. eximia and H. wrightorum are also closely related to H. euphorbiacea. Hyla regilla and H. cadaverina are more closely allied to each other than they are to the other three forms. A strong correlation (r = 0.93) between genetic relatedness as measured by immunological comparisons of albumin and that measured by electrophoretic studies of many proteins was found, although immunological techniques cannot be used to study taxa which are very closely related (D = 0.25 or less).