Eighty per cent of blood culture isolates of K-1 Escherichia coli are “resistant” to in vitro opsonization and phagocytosis by normal human leukocytes and serum. To investigate why most of bacteremic K-1 E. coli are poorly opsonized by normal human serum, we undertook this study to assess the serum opsonic requirements for K-1 isolates particularly in regard to the role of the alternative pathway of complement activation. To ascertain the role of complement in opsonization, we used an in vitro opsonic and bactericidal killing system and a method that determines complement activation by measuring consumption of hemolytic complement. For both assays, MgEGTA was used to specifically block the classic pathway while leaving the alternative intact. Additionally, we used the bactericidal killing assay to determine the possible participation of specific opsonins, like antibody, in the opsonization of these E. coli by using a serum source that was either previously absorbed with bacteria to remove antibody or by using purified K-1 capsular antigen to block specific K-1 antibody. Data demonstrate that E. coli K-1 are opsonized only by the classic pathway of complement via antibody that is not specific for the capsular K-1 antigen. Furthermore, the hemolytic complement consumption method was shown to be far less sensitive in determining which complement pathway is activated during opsonization of E. coli K-1. The restriction of these bacteremic K-1 E. coli isolates to the classic pathway of complement activation and the requirements for specific antibody for effective opsonization may be important factors underlying the pathogenicity of these bacteria.