FIG. 4.
Antibodies raised against the recombinant AS44–331 protein bind to the surface on E. faecalis cells only when Asc10 is expressed. Surface localization of the AS44–331 fragment was determined by flow cytometry analysis of intact E. faecalis OG1SSp(pMSP7517) cells. In experiments shown in panels A and C, Asc10 is not expressed. Expression of AS was induced (B and D) by the addition of 25 ng of nisin/ml to cultures. Panels A and B represent binding of a monoclonal antibody known to recognize surface-exposed Asc10, and panels C and D represent polyclonal antiserum raised against recombinant AS44–331. Secondary antibodies were labeled with fluorescein isothiocyanate. The percentage of gated populations is shown. Similar results were seen with E. faecalis OG1RF containing pMSP7517 (data not shown).