Murine models for bacterial superantigens like staphylococcal enterotoxin B (SEB) have to date been rather cumbersome. The reasons include: (1) necessary use of potentiating agents such as actinomycin D, d-galactosamine, lipopolysaccharide (LPS), or viruses; (2) high toxin amounts required to elicit effects; and/or (3) generation of phenotypic-stable transgenic animals. Our study employed readily available C3H/HeJ (TLR4 negative, LPS-nonresponsive) mice with intranasal and intraperitoneal administration of low microgram quantities of SEB. These animals responded to SEB with severe lung inflammation and hypothermia, culminating in death. A survey of cytokines/chemokines in sera and lungs after lethal intoxication revealed that monocyte chemoattractant protein-1 and interleukin-2 were associated with effects in this model. In contrast, SEB had minimal effects upon congenic (TLR4 positive, LPS-responsive) C3H/OuJ mice. Lethality of SEB in C3H/HeJ mice was neutralized with SEB-specific antibodies, suggesting potential utility of this model for future therapeutic studies.