Enhancing Polymyxin B Activity against Gram-Negative Bacteria Using Cubosomes

Quartz crystal microbalance with dissipation monitoring (QCM-D) was used to evaluate binding and interactions between biological molecules and surfaces in liquid. Experiments employed a Q-SENSE E4 system with an axial flow chamber, an IPC High Precision Peristaltic Multi-Channel Dispenser (ISMATEC, flow rate 0.2 mL/min), and 50 nm SiO2-coated quartz crystal sensors at 25 °C. Sensors were cleaned with 10 mM sodium dodecyl sulfate (SDS) for 8 h, rinsed extensively with Milli-Q water, dried under nitrogen, and oxidized in a UV-ozone chamber for 15 min to remove residual organic impurities. Upon adsorption of materials to the sensor surface, shifts in resonance frequency (Δf, reflecting mass of adsorbates including coupled water) and changes in energy dissipation (ΔD, correlating with viscoelastic properties of layers) were recorded using QSoft401 V2.5.2.418. Frequency and dissipation were measured simultaneously at 15 and 25 MHz, corresponding to the 3rd and 5th overtones of a 5 MHz crystal. Mass changes (Δm) were related to frequency changes via the Sauerbrey equation: Δm ≈ C−1/n * Δf, where n is the overtone number and C ≈ 17.7 ng/(cm2·Hz) for a 5 MHz AT-cut quartz crystal at room temperature. Thus, addition of 17.7 ng/cm2 of mass causes a 1 Hz frequency decrease; the 5 MHz quartz frequency can be measured with 0.01 Hz precision in vacuum, enabling nanogram-scale mass detection. For example, a monolayer of water (~25 ng/cm2) yields ~1.4 Hz frequency shift, within detection limits.