2.6. Synergistic Antibacterial Analysis

The synergistic antibacterial activity of the PMB-Se-NPs was assessed by comparison with those of the Se-NPs with PMB added at an equivalent dose. PI staining was utilized to evaluate bacterial viability and determine the degree of bacterial cell death, which could be quantified through flow cytometry. The bacterial suspension (1 × 10⁷ CFU mL⁻¹, 1 mL) was first incubated with varying concentrations of PMB-Se-NPs in a PBS solution (16, 32, and 64 μg mL⁻¹) for 10 min under light conditions (wavelength: 450 nm, irradiance: 3 mW cm⁻²). The experimental concentration of the nanoparticles was selected based on the minimum inhibitory concentration (MIC) results. The determination of the minimum inhibitory concentration (MIC) was conducted following the guidelines outlined in the CLSI Performance Standards for Antimicrobial Susceptibility Testing M100, 32nd edition. Untreated bacteria were used as the control. Subsequently, the suspension was centrifuged and the bacteria were resuspended in a PI solution (30 μM, 1 mL) to label dead bacteria. The fluorescence intensity of the PE channel was then measured using flow cytometry.