Bactericidal assay of Streptococcus gordonii strains treated with human polymorphonuclear leukocytes (PMN)

Streptococci were pre-incubated with 2% anti-Hsa antibody for 30 min at room temperature, thereafter that added 20% autologous serum (from the same donor from whom PMNs were isolated) was added and incubated for 30 min at room temperature, it was then washed, and resuspended in supplemented RPMI medium. Briefly, PMNs (5x106 cells) and the opsonized bacteria (1x106 cells) were suspended in 1 ml supplemented RPMI medium, mixed, and incubated in 1.5-mL tubes for 1 and 2 h at 37 °C with mixing using a rotator. The PMNs were disrupted with sterile water at room temperature, diluted with RPMI medium, and plated on BHI agar. Colonies were counted after incubation of the plates at 37 °C for 2 days.