Penicillin-binding proteins (PBPs) from penicillin-susceptible strains of Streptococcus pneumoniae are believed to be fairly similar in contrast to PBPs occurring in resistant isolates. The antigenic variation of PBPs la and 2b in 65 penicillin-susceptible strains from different geographic areas and a wide variety of isolation sites was analyzed using a set of specific antisera and monoclonal antibodies. Three strains contained different antigenic variants of PBP la, and 50 strains contained one of three antigenic variants found in PDP 2b. Seven patterns of antibody reactivity could be defined; all but one were distinct from those found recently in resistant strains. In addition, electrophoretic mobilities of all six PBPs, compared after conventional SDS-PAGE and fluorography, revealed an unexpected variation of PBP-profiles even for strains of one sero-group. Few strains appeared identical to each other or to the laboratory reference strain R6.