2.6. Assays for H. pylori Adhesion and Invasion to AGS Cells

Analyses of anti-adhesion and anti-invasion of H. pylori to AGS cells by Lactobacillus spp. were performed as previously reported [34]. Briefly, the live or heat-inactivated Lactobacillus strains suspension were added directly to the cell culture at an multiplicity of infection (MOI) of 100 for 10 min prior to inoculation with H. pylori 26695 (MOI = 100) to cells and incubation for 6 h. To determine the number of cell-adhesion bacteria, the infected cells were washed three times to remove unbound H. pylori and lysed with distilled water for 10 min. The lysates were then diluted in PBS and plated onto Brucella blood agar plates. After being cultured for 3–4 days, the viable CFUs were counted. To determine the number of intracellular H. pylori, the infected cells were washed three times in PBS followed by incubation with 100 μg/mL gentamicin (Sigma–Aldrich, Whitehouse Station, NJ, USA) for 1.5 h at 37 °C to remove extracellular bacteria. The following protocol was the same as above to obtain the viable CFUs. Results were determined by three independent experiments. The controls, containing H. pylori infected AGS cells, without Lactobacillus strains, were used to define as 100% adhesion or invasion. Results were expressed as the percentage of relative inhibition of H. pylori adhesion or invasion, by comparison with the controls, respectively.