BACKGROUND: Hemolytic disease of the fetus and newborn (HDFN) is a severe disease, resulting from maternal red cell (RBC) alloantibodies directed against fetal RBCs. The effect of a first‐trimester antibody screening program on the timely detection of HDFN caused by antibodies other than anti‐D was evaluated. STUDY DESIGN AND METHODS: Nationwide, all women (1,002 in 305,000 consecutive pregnancies during 18 months) with alloantibodies other than anti‐D, detected by a first‐trimester antibody screen, were included in a prospective index‐cohort study. In a parallel‐coverage validation study, patients with HDFN caused by antibodies other than anti‐D, that were missed by the screening program, were retrospectively identified. RESULTS: The prevalence of positive antibody screens at first‐trimester screening was 1,232 in 100,000; the prevalence of alloantibodies other than anti‐D was 328 in 100,000, of which 191 of 100,000 implied a risk for occurrence of HDFN because the father carried the antigen. Overall, severe HDFN, requiring intrauterine or postnatal (exchange) transfusions, occurred in 3.7 percent of fetuses at risk: for anti‐K in 11.6 percent; anti‐c in 8.5 percent; anti‐E in 1.1 percent; Rh antibodies other than anti‐c, anti‐D, or anti‐E in 3.8 percent; and for antibodies other than Rh antibodies or anti‐K, in none of the fetuses at risk. All affected children, where antibodies were detected, were promptly treated and healthy at the age of 1 year. The coverage validation study showed a sensitivity of the screening program of 75 percent. Five of 8 missed cases were caused by anti‐c, with delay‐induced permanent damage in at least 1. CONCLUSION: First‐trimester screening enables timely treatment of HDFN caused by antibodies other than anti‐D, however, with a sensitivity of only 75 percent. A second screening at Week 30 of c– women will enhance the screening program. Severe HDFN, caused by antibodies other than anti‐D, is associated with anti‐K, anti‐c, and to a lesser extent with other Rh‐alloantibodies.