Dentin Infiltration and Bacterial Viability Assay

Two microliters of S. mutans suspension at 10¹⁰ CFU per mL was placed on a dentin block to infiltrate into dentin, as described above. Then a primer was applied to the dentin and left for 20 s. Two microliters of primer was used for each dentin block. Six dentin blocks were used to test each primer (n=6). The dentin blocks were then stained using a live/dead bacterial viability kit (Molecular Probes, Eugene, OR, USA) following the manufacturer's protocol. Live bacteria were stained with Syto 9 to show a green fluorescence. Bacteria with compromised membranes were stained with propidium iodide to show a red fluorescence. The stained samples were imaged via an epifluorescence microscope (Eclipse TE2000-S; Nikon, Melville, NY, USA).^{37 (link), 42 (link)}