Stock solutions of gentamicin (ITW Reagents, Monza, Italy) were prepared at 4760 µg/mL, clindamycin (Cayman Chemical, Michigan, MI, USA) at 660 µg/mL, and vancomycin (ITW Reagents, Monza, Italy) at 1062 µg/mL, according to the manufacturer’s specifications, and kept at −80 °C. Antibiotic working solutions were used at their minimum inhibitory concentration (MIC) values (gentamicin, 0.238 µg/mL; clindamycin, 0.033 µg/mL; and vancomycin, 0.531 µg/mL), as determined in a previous study [65 (link)].
Nisin A (1000 UI/mg, 2.5% purity) (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in 0.02 M HCl (Merck, Darmstadt, Germany), yielding a 1000 µg/mL stock solution, and pexiganan (>95% purity; Innovagen, Lund, Sweden) was dissolved in deionized sterile water, yielding a 2048 µg/mL stock solution. These solutions were then filtered through a 0.22 µm filter (Millipore Corporation, Billerica, MA, USA) and kept at 4 °C. To prepare the dual-AMP biogel, a pexiganan solution at 256 µg/mL was enriched with Nisin At 125 µg/mL, corresponding to their minimum biofilm eradication concentration (MBEC) values, as determined previously [25 (link)]. 0.75 g of guar gum (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in 50 mL of sterile distilled water, yielding a 1.5% (w/v) gel, which was sterilized by autoclave. Antibiotics and AMPs were mixed into the gel in a 1:1 ratio.
Nisin A (1000 UI/mg, 2.5% purity) (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in 0.02 M HCl (Merck, Darmstadt, Germany), yielding a 1000 µg/mL stock solution, and pexiganan (>95% purity; Innovagen, Lund, Sweden) was dissolved in deionized sterile water, yielding a 2048 µg/mL stock solution. These solutions were then filtered through a 0.22 µm filter (Millipore Corporation, Billerica, MA, USA) and kept at 4 °C. To prepare the dual-AMP biogel, a pexiganan solution at 256 µg/mL was enriched with Nisin At 125 µg/mL, corresponding to their minimum biofilm eradication concentration (MBEC) values, as determined previously [25 (link)]. 0.75 g of guar gum (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in 50 mL of sterile distilled water, yielding a 1.5% (w/v) gel, which was sterilized by autoclave. Antibiotics and AMPs were mixed into the gel in a 1:1 ratio.
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