Lipid coated liquid crystal droplets for the on-chip detection of antimicrobial peptides

Lipid coated liquid crystal droplets have been trapped in a novel trap structure for the on-chip detection of a model antimicrobial peptide – Smp43, an α-helical peptide from Scorpion Venom.

The liposome solution was made by hydration and tip-sonication of dried lipid mixture (DOPC&DOPG 1:1 with 0.1mol% Texas Red-DHPE) in 10mM HEPES buffer (pH7.5), as described previously [ 1 ]. The Texas Red-DHPE was included to allow fluorescence imaging of the lipid-coated droplets, providing confirmation of the presence or removal of the lipid. The lipid mixture was dissolved in chloroform in a glass vial and dried under nitrogen flow for 1h. Buffer solution was then added and the vial was vortex-mixed for 2 minutes to produce a suspension of the lipids. The suspension is tip-sonicated for 30 min at 4 o C until it is clear. The resulting liposomes have a size of ~ 25-30 nm [ 1 ]. The liposome solution was normally kept at 4 o C and used within one week. 15 % volume of glycerol was added to the liposome solution to increase the viscosity, optimizing the flow properties for microfluidic liquid crystal droplet production.

Microfluidic device fabrication
To make the polydimethylsiloxane (PDMS) devices, a silicon master with a SU8 pattern was first fabricated. The SU8 patterns with a thickness of 25 μm were formed on a threeinch silicon wafer using a MW2 laser direct writing system (Durham Magneto Optics Ltd, Durham, UK). The SU8 pattern was then hard-baked at 210 o C for 15 minutes to improve the mechanical properties of the SU8 master. To make an inverse PDMS copy of the SU8/Si master, the PDMS monomer was first fully mixed with the curing agent at a weight ratio of 10:1. The master was placed at the bottom of a petri dish and the PDMS monomer mixture poured onto it to yield a thickness of ~5 mm. The system was then placed under vacuum for 30 minutes to remove any air bubbles. Finally, to crosslink the PDMS, the petri dish was placed in an oven at 75 o C for 1 hour.
The PMDS layer was temporarily transferred to a glass plate and holes at the inlets and outlets made using a biopsy punch. The PDMS device was plasma cleaned (100W, O 2 pressure 0.5mbar, 1min, Zepto Plasma Unit, Diener Electronic, Germany) and bonded to a second cleaned glass plate. Applying a gentle pressure and baking in an oven at 75 o C for 30min forms a PDMS microfluidic device that was ready for use.

Lipid-coated liquid crystal droplet production
Monodisperse lipid-coated droplets (diameter = 17 μm) were produced using a flow focus droplet microfluidic device [ 2 ]. A schematic diagram of the droplet formation process in the device is shown in Figure 2(a). The droplet formation device had two inlets. One inlet fed the two outer side channels, with buffer solution containing lipid in the form of small unilamellar liposomes. The middle inlet was used for the feeding of liquid crystal (E7). The E7 and liposome solutions were pumped into the device through the two inlets using two PHD ULTRA advanced syringe pumps (Harvard Apparatus, USA). The flow rate used for LC droplet generation was 0.075 μL/min for E7 and 10 μL/min for buffer with liposomes. Figure S1. Images of (a) the as-produced PC/PG (1:1) coated E7 droplets and (b) the same droplets experience a thermal treatment at 75 o C for 20min, taken under crossed polarizer in transmission mode of polarized microscope.    Figure S4 is bigger than that in Figure  3b because the size is determined from analysis of the high-speed camera images, which have a lower resolution than the still images used in Figure 3.

Support information 4 -Video:
Video S1. The formation of the PC/PG (1:1) coated E7 droplets using a microfluidic device, as mentioned in Figure2. Frame rate: 50k per second.
Video S2. The switch of the PC/PG (1:1) coated E7 droplets in the trap structure under the constant flow of SMP43. Images were taken every 5 min.