A rubella-specific lymphocyte transformation assay, using cryopreserved mononuclear cells, has been developed and used to evaluate specific responses among 21 children with congenitally acquired rubella (CAR), 25 healthy control children and 10 children with sensorineural deafness of unknown aetiology. Although all 21 children with CAR were seropositive, 12 (57.1%) failed to respond to rubella antigen in the transformation assay. Negative in vitro lymphocyte transformation responses were detected significantly more frequently among congenitally infected children below 3 years of age. Thirteen of the 25 (52%) control children were seropositive; only one of these seropositive children (7.6%) gave a negative transformation response. A negative rubella-specific lymphocyte transformation response in a seropositive child, particularly when aged 3 years or younger, is therefore suggestive of CAR. Four of the 10 children with deafness of unknown aetiology were rubella seropositive but gave negative responses in the transformation assay, suggesting that these children had CAR. Our assay may provide a very useful test for retrospective diagnosis of CAR, particularly in children under the age of 3.