Fig. 5: Correlation between the number of live bacteria (S. aureus) and the band-dependent AMP density on the gradient Ti surfaces and the use of information from such correlation to prepare nongradient surfaces for optimized bacteria killing.

a Distribution of live bacteria (red dots, detected by Petrifilm method) on different bands of Ti-Grad-20AMP after 24 h of culturing, showing that higher band number area (bands 7–10, with higher AMP densities) did not have any live bacteria, whereas lower band number area (bands 1–6, with lower AMP densities) presented live bacteria. Three lines were denoted, including P1, P2, and P3, corresponding to the boundary between the area without and with dead bacteria, starting boundary of band 7, and midpoint position within band 7, respectively (n = 3, scale bar, 1.5 mm). b FITC fluorescence images of the nongradient Ti surface prepared using the conditions (immersion time and AMP concentration during immersion) corresponding to P1, P2, and P3 in a, producing Ti-AMP-P1, Ti-AMP-P2, and Ti-AMP-P3, respectively. For each sample, five images were chosen randomly to calculate the MFI (n = 5, scale bar, 500 μm). c Bacterial viability of Ti–S, AMP-P1, Ti-AMP-P2, and Ti-AMP-P3 (n = 3) (Sidak’s multiple comparisons test, two-way ANOVA. Ti–S vs Ti-AMP-P1, **p < 0.0001; Ti–S vs Ti-AMP-P2, **p < 0.0001; Ti–S vs Ti-AMP-P3, **p < 0.0001). d CCK-8 results for the indicated uniform Ti surface with mBMSCs after 1 and 3 days of culturing (n = 3). Ti–S was the Ti surface modified with silane-PEG2000-MAL. Data are displayed as mean ± SD and analyzed by GraphPad Prism software.