Fig. 2. Host-cell-associated multimeric vitronectin promotes adherence to and invasion of pneumococci into host tissue cells. (A) Pneumococcal (NCTC10319) adherence (at 4 hours) was investigated by immunofluorescence staining after pre-incubation of Detroit 562 cells with 3.0 µg native vitronectin or 3.0 µg of commercially purchased (com) or 3.0 µg self-generated (gen) multimeric vitronectin. The number of attached bacteria per cell in the absence of vitronectin (none) was used as control. Results represent the mean ± s.d. of at least three independent experiments, each done in duplicate. ^*P<0.005; n.s., non-significant. (B) Multimeric vitronectin promotes the adherence of pneumococci to Detroit 562 cells in a dose-dependent manner. (C) Adherence of pneumococci to the human epithelial cells Detroit 562 or A549, and the endothelial cell line HBMEC after 4 hours of infection was determined by immunofluorescence in the absence (none) or presence of 3.0 µg multimeric vitronectin (VN). Results represent the mean ± s.d. of at least three independent experiments, each done in duplicate. ^*P<0.05. (D) Immunofluorescence microscopy of pneumococci attached to host cells in the absence (none) or presence of multimeric vitronectin. (E) Invasion and intracellular survival of pneumococci in Detroit 562 epithelial cells and the endothelial cell line HBMEC in the absence (none) or presence of multimeric vitronectin (VN) was determined by the antibiotic protection assay. Results represent the mean ± s.d. of at least three independent experiments, each done in duplicate. ^*P<0.03.