Immunoelectron microscopic studies of Actinomyces viscosus

Optimal conditions for labelling Actinomyces viscosus (T14V) in thin sections by an indirect immunoperoxidase technique are described. The influence of several fixatives on the degree of labelling of bacterial cells was tested in pure cultures and in dental plaques. Some pure cultures were also processed without prior fixation. The effect of different incubation conditions on the degree of labelling was evaluated in pure cultures. The optimal labelling conditions included fixation with paraformaldehyde (4%) plus glutaraldehyde (0.25%), incubation with highly diluted rabbit anti‐T14V γglobulin for 24 hours at 4°C, followed by incubation with diluted sheep anti‐rabbit IgG conjugated to horseradish peroxidase, and subsequent histochemical visualization of the peroxidase. Optimal labelling conditions were used to check for cross‐reactions between the anti‐T14V γ‐globulin and some common oral bacteria. The well‐known cross‐reaction with Actinomyces naeslundii was noted, as well as a slight cross‐reaction with Bacterionema matruchotii. Optimal conditions were used for labelling sections of dental plaques. Wide variations were observed in the number of labelled cells. Cells labelled with anti‐T14V γ‐globulin were unlabelled when consecutive sections of these cells were treated with control normal rabbit serum γ‐globulin or anti‐Streptococcus mutansγglobulin. Cells labelled with anti T14V γglobulin tended to be cocco‐bacillary in shape in the superficial layers of plaque. The deeper layers exhibited more filamentous forms. These observations are compatible with the known pleomorphism of A. viscosus in vitro.