Previous results have demonstrated that nonspecific protective immunity against lethal Francisella tularensis live vaccine strain (LVS) or Listeria monocytogenes infection can be stimulated either by sublethal infection with bacteria or by treatment with bacterial DNA given 3 days before lethal challenge. Here we characterize the ability of purified lipopolysaccharide (LPS) from F. tularensis LVS to stimulate similar early protective immunity. Treatment of mice with surprisingly small amounts of LVS LPS resulted in very strong and long-lived protection against lethal LVS challenge within 2 to 3 days. Despite this strong protective response, LPS purified from F. tularensis LVS did not activate murine B cells for proliferation or polyclonal immunoglobulin secretion, nor did it activate murine splenocytes for secretion of interleukin-4 (IL-4), IL-6, IL-12, or gamma interferon (IFN-gamma). Immunization of mice with purified LVS LPS induced a weak specific anti-LPS immunoglobulin M (IgM) response and very little IgG; however, infection of mice with LVS bacteria resulted in vigorous IgM and IgG, particularly IgG2a, anti-LPS antibody responses. Studies using various immunodeficient mouse strains, including LPS-hyporesponsive C3H/HeJ mice, muMT(-) (B-cell-deficient) knockout mice, and IFN-gamma-deficient mice, demonstrated that the mechanism of protection does not involve recognition through the Lps(n) gene product; nonetheless, protection was dependent on B cells as well as IFN-gamma.