Fluorescence microscopy.

The binding specificity of GPB to various bacterial strains was examined by fluorescence microscopy after incubation with an EGFP-fused GPB protein, EGFP-GPB (EGPB). Briefly, various bacterial strains, including S. pneumoniae NS26, S. pneumoniae NS63, S. agalactiae, S. dysgalactiae, S. pyogenes, and S. mutans, were grown overnight, washed and suspended in PBS to an OD₆₀₀ of 0.5 to 0.8. Next, 100 μl of each bacterial suspension was incubated with an equal concentration (20 μM) of EGPB or EGFP protein for 60 min at 37°C. After three washes with PBS, the samples were imaged by a Nikon fluorescence microscope (TS2-FL) at ×100 magnification and with a 0.4-s exposure time.