Fig 2.

A dot blot amylase overlay assay was used to demonstrate amylase binding to bacterial cells. Whole bacterial cells were pelleted from broth culture, washed, and standardized to an OD₆₀₀ of 2.9 in PBS. Each sample (10 μl) was spotted onto a nitrocellulose membrane, blocked with skim milk, overlaid with salivary amylase (0.1 mg/ml) followed by a rabbit anti-amylase antibody, and developed with alkaline phosphatase-conjugated goat anti-rabbit IgG and SigmaFAST BCIP-NBT (Sigma). CH1, S. gordonii Challis CH1; AbpA⁻, the KS1 amylase-binding protein A mutant; AbpB⁻, the KS1 amylase-binding protein B mutant.