Genetic variation, using blood groups and red cell and serum proteins, was surveyed in the Cabecar of Chirripo and the Huetar of Quitirrisi, Costa Rica. Thirty-nine loci were screened in a sample of 91 Cabecars and 40 loci in 45 Huetars. Twenty-seven loci were monomorphic in the Cabecar and 30 in the Huetar. The proportions of polymorphic loci (P), out of 34 studied by electrophoresis, were 0.235 and 0.177, respectively. Estimated gene diversities (H) of the polymorphic loci were 0.050 in the Cabecar and 0.053 in the Huetar. Two polymorphisms, reported until now in Costa Rican and Panamanian Chibchan groups only, occurred at very high frequencies: TF*DGUA = 0.357 in the Cabecar, the highest frequency ever reported, and 0.033 in the Huetar; and PEPA*F, which reached 0.26 in the Cabecar and 0.29 in the Huetar. Nei's genetic distances and trees (two methods) were used to compare them to seven other Chibchan tribes of Costa Rica. The results placed both the Cabecar of Chirripo and the Huetar closer to the Talamancan Tribes (Bribri and Cabecar). This was an unexpected result for the Huetar, since linguistic studies suggested a closer relationship to the Guatuso. GST, DST, RST, and Dm for three Cabecar subpopulations (Atlantic, Chirripo, and Pacific) doubled their values compared to estimates based on comparison of only two subpopulations: Atlantic and Pacific. Total genetic diversity considering just the three Cabecar subpopulations resembled that obtained including them plus six other Chibchan populations of Costa Rica.