The risk of contamination of tissue culture cells with transmissible spongiform encephalopathy (TSE) agents as a result of the use of animal products as medium components has been considered to be low, in part, because only a few brain-derived cell lines have been reported to be susceptible to TSE infection. In the present study, we demonstrate that the common laboratory fibroblast cell lines NIH/3T3 and L929, which express low levels of cellular mouse prion protein, are susceptible to infection with mouse-adapted scrapie. Our results show that the susceptibility of a cell line to TSE infection cannot be predicted on the basis of its tissue origin or its level of expression of the cellular prion protein, and they suggest that any cell line expressing normal host prion protein could have the potential to support propagation of TSE agents. Thus, testing of cells for TSE susceptibility might be necessary for all cell lines that are routinely used in vaccine production and in other medical applications.

Vaccination with replication-competent vaccinia protects against heterologous orthopoxvirus challenge. CD4 T cells have essential roles helping functionally important Ab and CD8 antiviral responses, and contribute to the durability of vaccinia-specific memory. Little is known about the specificity, diversity, or dominance hierarchy of orthopoxvirus-specific CD4 T cell responses. We interrogated vaccinia-reactive CD4 in vitro T cell lines with vaccinia protein fragments expressed from an unbiased genomic library, and also with a panel of membrane proteins. CD4 T cells from three primary vaccinees reacted with 44 separate antigenic regions in 35 vaccinia proteins, recognizing 8 to 20 proteins per person. The integrated responses to the Ags that we defined accounted for 49 to 81% of the CD4 reactivity to whole vaccinia Ag. Individual dominant Ags drove up to 30% of the total response. The gene F11L-encoded protein was immunodominant in two of three subjects and is fragmented in a replication-incompetent vaccine candidate. The presence of protein in virions was strongly associated with CD4 antigenicity. These findings are consistent with models in which exogenous Ag drives CD4 immunodominance, and provides tools to investigate the relationship between Ab and CD4 T cell specificity for complex pathogens.

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