The ability of heat-killed Mycobacterium vaccae to stimulate a cytotoxic T-cell response to an unrelated protein is associated with a 65 kilodalton heat-shock protein.

SUMMARY Exogenous antigens are generally presented by Class II major histocompatibility (MHC) molecules. When administered with an adjuvant, however, they are capable of inducing a CD8 + T-cell response where antigen recognition is associated with Class I MHC. Accordingly, immunization with soluble ovalbumin (OVA) alone does not activate CD8 + cytotoxic T cells (CTL) but when given in complete Freund’s adjuvant (CFA), or in formulations of a number of novel adjuvants, an OVA-specific CD8 + CTL response can be detected. We show in this report that immunization with soluble OVA mixed with heat-killed Mycobacterium vaccae, but not with other common pathogenic and saprophytic mycobacteria, can activate OVA-specific CD8 + CTL. An OVA-specific CTL response is detected when mice are immunized by either the intraperitoneal or intranasal route and their spleen cells are re-stimulated in vitro. Adjuvant activity of heat-killed M. vaccae is present in M. vaccae culture filtrate, in soluble protein components of whole M. vaccae and in the 65 kDa heat-shock protein (hsp) of M. vaccae. Mycobacterium vaccae has previously been shown to have no adverse side-effects in humans. The current results suggest that M. vaccae may be useful as an adjuvant for vaccines and other immunotherapies where CD8 + CTL responses to exogenous