The biochemical defect that underlies the genetic disorder cystic fibrosis (CF) has been proposed to involve an altered regulation of epithelial Cl- permeability by agents such as adenosine 3',5'-cyclic monophosphate (cAMP). We report here the successful complementation of this functional defect achieved by using the technique of somatic cell fusion to introduce the normal CF allele into mutant cells. CF epithelial cells were fused with transfectant mouse fibroblasts that contain the normal human gene. The resulting heterokaryons were examined for restoration of cAMP-activated Cl- transport using an optical assay of Cl- permeability. Our results provide direct evidence for the involvement of the protein product of the normal CF allele in modulating epithelial Cl- permeability.