Immune phage antibody libraries are an attractive technology for isolating antigen-specific monoclonal antibodies (mAbs). Here we show that the immunization schedule affects the immune phage antibody library properties. We subcutaneously (s.c.) administered HIV-1 Nef and Vif antigens with different schedules (25 mu g x 2 s.c. and 10 mu g x 3 s.c.). The variety of isolated mAbs in 25 mu g x 2 s.c. groups (Nef: 11 clones, Vif: 9 clones) was superior to that in the 10 mu g x 3 s.c. groups (Nef: 2 clones, Vif: 1 clone). This finding suggests that it is important to optimize the immunization schedule for isolating a wide variety of mAbs. (C) 2011, The Society for Biotechnology, Japan. All rights reserved.