The aim of the present investigation was to test a procedure useful for the electron-microscopic in situ identification of the presumptive cariogenic microorganism Streptococcus mutans (serotype d) growing in the human dental plaque. For this purpose, different parameters of an indirect immunohistochemical method were tested in three sets of experiments. In experiment set I, all serological and histochemical procedures were performed en bloc on specimens fixed only with glutaraldehyde before embedding in Vestopal W. Different marking substances, such as ferritin, alkaline phosphatase and horseradish peroxidase (HRP) were tested. The en bloc method using HRP-labelled antibodies was found useful for staining of in vitro-grown bacteria but failed when applied to dental plaque. In experiment set II, ultrathin sections of glutaraldehyde-fixed and glycol methacrylate-embedded in vitro-grown bacteria were section-stained. The bacteria became outlined with a highly electron-dense HRP reaction product which accumulated on top of the cell envelope. The same type of HRP reaction product was found on some bacteria in the 2-day-old dental plaque after section-staining. This system was further tested in a series of controls also using consecutive serial sections. In exp. set III, a number of different stationary and transient oral bacteria were immunohistochemically section-stained. A cross-reaction with bacteria belonging to S.salivarius was discovered and removed by absorbing the anti-S.mutans serotype d serum with S.salivarius (NCTC 8618).
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