Chloroform stocks of d‐DMPC and d‐POPC were used to prepare the lipid films. Lipid films were formed by removing chloroform through evaporation under a flow of nitrogen, and any residual organic solvent was eliminated under vacuum. The films were stored at −20°C until use. The vesicle fusion method was used to form the SLBs, according to previous protocols (Lind et al., 2014 (link)). These films were hydrated before use in a bath sonicator, at 40–50°C, for 1 h in either MilliQ water for NR experiments or D2O for ATR‐FTIR. Immediately before deposition, the lipids were tip sonicated (5 min on and 5 min off) for 15 min at 20% power (400 W, Branson 450 Digital Sonifier w/Probe), mixed with CaCl2 to a final concentration of 2 mM, and injected into the flow cell at a lipid concentration of 0.1 mg/mL. The lipids were incubated for 20 min before rinsing with MilliQ water or D2O followed by h‐TBS or d‐TBS, for NR or ATR‐FTIR, respectively.
450 digital sonifier w probe
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The 450 Digital Sonifier w/Probe is a lab equipment product that provides ultrasonic homogenization and disruption of samples. It is designed to operate at a specific frequency and power output to achieve efficient sample processing.
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2 protocols using 450 digital sonifier w probe
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Formation of Supported Lipid Bilayers
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Supported Lipid Bilayer Formation
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Lipid films were prepared by dissolving the deuterated phospholipids and d-cholesterol in chloroform and mixing at the desired molar ratio (i.e., 80:20 mol% phospholipid: cholesterol), followed by evaporation of the organic solvent under nitrogen gas. Once the film was formed any residual chloroform was removed by leaving the films under a vacuum overnight and then stored at −20 °C until needed.
SLBs were prepared via a small unilamellar vesicle fusion method following previously reported protocols [26] (link). First, multilamellar vesicles were formed by hydrating lipid films with Milli-Q water and the mixture was kept in a bath sonicator at a temperature between 40 and 50 °C for 1 h. Second, the suspension was tip sonicated (5 min on and 5 min off) for 15 min at 20% power (400 Watts, Branson 450 Digital Sonifier w/ Probe). Before injection, 0.2 mg/mL lipid vesicles were mixed with an equal volume of 4 mM Calcium Chloride (CaCl2). The vesicles were introduced into the cells by a pump at 1 mL/min and incubated for 20 min. This was followed by extensive rinsing with Milli-Q water and h-TBS buffer (TBS in H2O).
SLBs were prepared via a small unilamellar vesicle fusion method following previously reported protocols [26] (link). First, multilamellar vesicles were formed by hydrating lipid films with Milli-Q water and the mixture was kept in a bath sonicator at a temperature between 40 and 50 °C for 1 h. Second, the suspension was tip sonicated (5 min on and 5 min off) for 15 min at 20% power (400 Watts, Branson 450 Digital Sonifier w/ Probe). Before injection, 0.2 mg/mL lipid vesicles were mixed with an equal volume of 4 mM Calcium Chloride (CaCl2). The vesicles were introduced into the cells by a pump at 1 mL/min and incubated for 20 min. This was followed by extensive rinsing with Milli-Q water and h-TBS buffer (TBS in H2O).
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