A simple and reproducible method is described for the measurement of proliferative responses of peripheral blood lymphocytes in whole blood upon stimulation with horse anti-human lymphocyte serum (ALS), phytohaemagglutinin (PHA) or a monoclonal antibody (mAb) directed against CD3. Only small aliquots of blood are needed and separation procedures are not necessary. Since the method proved to be very reproducible and showed little variation, reference values of proliferation, expressed in cpm per 15 microliters blood and cpm per 10(3) CD3+ lymphocytes, were determined based upon the 5th and the 2.5th percentile of the distribution of the values in our reference population. In addition, the use of whole blood lymphocyte cultures in longitudinal studies of immunocompromised individuals is demonstrated.