Identification of Actinobacillus actinomycetemcomitans by gold immunolabeling and scanning electron microscopy

Recent evidence suggests that dental plaque is not a homogeneous bacterial mass but, on the contrary, specific bacterial morphotypes and species may be preferentially located within certain microenvironments. The aim of the present study was to develop combined gold immunolabeling and scanning electron microscopic (SEM) techniques for the identification of periodontal pathogens in subgingival dental plaque and on the root surfaces of extracted teeth. Suspensions of pure A. actinomycetemcomitans cultures or suspensions of A. actinomycetemcomitans mixed together with other oral bacteria were prepared, labeled with goat anti-rabbit IgG conjugated with 5 nm or 40 nm colloidal gold particles, and observed by SEM using both secondary and back-scattered imaging. Scanning electron microscopy demonstrated that the A. actinomycetemcomitans bacterial cell surface was specifically labelled. There was no cross-reaction with any of the other bacterial morphotypes. Only the labeled A. actinomycetemcomitans were visible in mixtures examined by back-scattered imaging scanning electron microscopy. The combined techniques of gold immunolabeling and SEM may, therefore, be useful in identifying A. actinomycetemcomitans in subgingival plaque samples and on the root surfaces of extracted teeth as well as in studies of bacterial ecology in dental plaque, in general.