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4 protocols using 1 palmitoyl 2 oleoyl sn glycero 3 phosphocholine

1

Lipid Preparation for Membrane Studies

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1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1, 2- dioleoyl-sn-glycero-3-phosphocholine (DOPC) were purchased from the NOF Corporation (Tokyo, Japan). 1,2-dioleoyl-sn-glycero-3-phsphoEthanolamine-N-(lissamine rhodamine B sulfonyl) (Rho-PE) was purchased from Avanti Polar Lipids, Inc. (810150C, Alabaster, AL, USA). Ethanol, sodium chloride, and chloroform were purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan).
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2

Lipid Nanoparticle Formulation Protocol

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1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-dioleoyl-sn-glycero-3-phosphoEthanolamine (DOPE), 1,2-dioleoyl-3-dimethylammonium-propane (DODAP), and 1,2-dimyristoyl-rac-glycero-3-methoxypolyethylene glycol-2000 (DMG-PEG2K) were purchased from the NOF Corporation (Tokyo, Japan). Cholesterol was purchased from Sigma-Aldrich (St. Louis, MO, USA). A pH-sensitive cationic lipid, CL4H6, was synthesized in the laboratory as previously described.19 (link) Ethanol, sodium chloride, citric acid, 2-morpholinoethanesulfonic acid (MES), monohydrate, and phosphate-buffered saline (PBS) were purchased from FUJIFILM Wako Pure Chemical Corporation (Osaka, Japan). The siFVII and Cy-5-labeled siGL4 were purchased from Hokkaido System Science Co. Ltd. (Sapporo, Japan). Table S4 shows the sense and antisense strand sequences of siFVII and Cy5-siGL4. Quant-iT™ Ribogreen™ RNA reagent and 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate (DiI) were obtained from Thermo Fisher Scientific (Waltman, MA, USA). SU-8 3050 was obtained from Nippon Kayaku Co., Ltd. (Tokyo, Japan).
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3

Lipid Membrane Preparation Protocol

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1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), 1,2-dio-leoyl-sn-glycero-3-phosphocholine (DOPC), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE) were procured from NOF Corporation (Tokyo, Japan). 1-Palmitoyl-2-oleoyl-sn-glycero-3-phospho-L-serine (POPS) was procured from Avanti Polar Lipids (Alabaster, AL, USA). 1-Palmitoyl-2-(1-pyrenedecanoyl)-sn-glycero-3-phosphocholine (py-PC) was purchased from Molecular Probes (Eugene, OR, USA). Ergosterol was acquired from Sigma-Aldrich Corporation (St. Louis, MO, USA).
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4

Preparation of POPC/POPG/Chol LUVs

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1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1’-rac-glycerol) (POPG) were obtained from NOF Corporation. For NBD binding assays, a lipid film was formed by rotary evaporation of a mixture of POPC/POPG/Chol (molar ratio = 4:1:2) in chloroform. After vacuum-drying overnight, the lipid film was hydrated with phosphate buffered saline (PBS: 137 mm NaCl, 2.7 mm KCl, 10 mm Na2HPO4, and 1.8 mm KH2PO4. LUVs were generated through 21 successive extrusions of the lipid dispersions through double polycarbonate filters with 100-nm pore size, using a Liposofast extrusion system (Avestin). The concentration of LUVs was determined using a LabAssay Phospholipid kit (Wako) and expressed as the lipid concentration.
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