A plaque assay for assessment of immune responses to Streptococcus mutans cell wall carbohydrate

A plaque-forming cell (PFC) assay has been developed for measurement of single cell responses to the serotype carbohydrate antigen of the Streptococcus mutans cell wall. Serotype g carbohydrate was purified from a mutanolysin (M1) enzyme digest (and designated M1g) of S. mutans 6715 cell walls by ion exchange and gel filtration chromatography. M1g carbohydrate was esterified by stearoylization (sM1g), and subsequently coated to sheep erythrocytes (sM1g-SRBC). This coating antigen was then used for enumeration of IgM anti-M1g PFC responses from spleens of either mice or rats immunized with S. mutans 6715 antigen. Good splenic IgM anti-M1g PFC responses were seen in either mice or rats given S. mutans whole cells or cell walls, while cell wall lysates or M1g were lowly immunogenic. Of interest was the finding that sM1g induced good IgM anti-M1g PFC responses in mice and in murine spleen cell cultures, in vitro. This study describes a method for assessment of individual antibody-producing cells to a major S. mutans cell wall determinant which should facilitate studies directed to determine mechanisms involved in the induction of immune responses to this important bacterium.