Murine monoclonal subunit-specific antibodies against heterooctameric phosphofructokinase-1 from Saccharomyces cerevisiae exhibiting no cross-reactivity were purified and characterized regarding complex formation with the native enzyme, immunological reactivity to the SDS-denatured subunits originating from native and proteolytically truncated phosphofructokinase, and protection against proteolytic degradation. Strong complex formation was found with one α-specific antibody. Western blotting employing different enzyme forms allowed to localize epitope sites of the α-polypeptide chain. Monoclonal antibodies protect phosphofructokinase-1 against chymotryptic degradation.