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The interaction of host cells with microbial products or their invasion by pathogens frequently results in activation of the NF-κB family of transcription factors. The studies presented here reveal that in vivo, infection with Toxoplasma gondii results in the activation of NF-κB. To determine whether host cells could activate NF-κB in response to invasion by T. gondii, Western blots, immunofluorescence, and electrophoretic mobility shift assays were used to assess the response of host cells to infection. In these studies, infection of macrophages or fibroblasts with T. gondii did not result in the activation of NF-κB. In addition, the ability of lipopolysaccharide to activate NF-κB was impaired in cultures of macrophages infected with T. gondii. Together, these data demonstrate that invasion of cells by T. gondii does not lead to the activation of NF-κB and suggest that the parasite may actively interfere with the pathways that lead to NF-κB activation.