OBJECTIVE. To establish the prevalence of human T-lymphotropic virus (HTLV) type I among Dutch blood donors. DESIGN. Study to confirm screening results. SETTING. Central Laboratory of the Blood Transfusion Service of the Dutch Red Cross, section Virus Diagnostics, Amsterdam. METHOD. The majority (550,000) of the Dutch blood donors were tested in the course of 1993 for presence of HTLV antibodies. For serological confirmation using the Western Blot test, 20 of the 22 Dutch blood banks sent in blood samples of 714 donors found positive at HTLV screening. Material of 36 of these donors was suitable for testing using the polymerase chain reaction (PCR) for DNA of HTLV types I and II. RESULTS. The Western Blot confirmation test confirmed HTLV infection of 10 of the 714 blood donors; eight of these samples, examined with the PCR test, proved to be positive for DNA of HTLV I. In 537 of the 714 ELISA-reactive donors, the Western Blot test gave doubtful results. The PCR test for HTLV I/II DNA was negative in 26 of these samples, but the possibility could not be excluded that there were HTLV carriers among the other donors with a doubtful Western blot test result. With inclusion of the four donors infected with HTLV I that had been detected earlier, the prevalence of HTLV I among Dutch blood donors is found to be 1:39,000.