The acidic drug (S)-Naproxen (Mw 230,26 g/mol, pka = 4.15, solubility in water 15.9 mg/L) was purchased from Fagron (Barsbüttel, Germany) and the basic amino acids L-arginine (MW = 174.2 g/mol, pka = 9.04 and 12.48) and L-lysine (MW = 146.2 g/mol, pka = 8.95 and 10.53) were purchased from Sigma Aldrich (St. Louis, MO, USA), respectively. The surfactants sodium dodecyl sulfate (MW ∼ 288 g/mol), pluronic F-127 (MW ∼ 12,600 g/mol) and polyoxyethylene (40) stearate (MW ∼ 328.5 g/mol) were also purchased from Sigma Aldrich (St. Louis, MO, USA). Tween 20 (MW ∼ 1227 g/mol) was donated by Merck (Darmstadt, Germany), while the TPGS 1000 (MW ∼ 1513 g/mol) was bought from Isochem (Vert-le-Petit, France). All substances were used as received. All surfactants are considered non-toxic in the used amount. The toxicity was considered based on Pub Chem, the National Library of Medicine, with the following LD 50 values: SDS (rat, oral: 1288 mg/kg), Tween 20 (rat, oral: 37 g/kg), pluronic (rat, oral: 22.4 g/kg) and polyoxyethylene stearate (mouse, i.p. 200 mg/kg). No toxicity data were available for TPGS.
Polyoxyethylene 40 stearate
Manufactured by Merck Group
Sourced in United States
Polyoxyethylene (40) stearate is a non-ionic surfactant used as an emulsifier and wetting agent in various pharmaceutical, cosmetic, and industrial applications. It has an average of 40 ethylene oxide units per molecule.
Automatically generated - may contain errors
Lab products found in correlation
1 2 distearoyl sn glycero 3 phosphocholine, by Avanti Polar Lipids (4 mentions)
Distearoylphosphatidylcholine, by Avanti Polar Lipids (3 mentions)
1 2 distearoyl sn glycero 3 phosphoethanolamine n methoxy polyethylene glycol 2000, by Avanti Polar Lipids (2 mentions)
Distearoyl phosphatidylserine, by Avanti Polar Lipids (2 mentions)
Streptomycin, by Thermo Fisher Scientific (2 mentions)
L arginine, by Merck Group (1 mentions)
Sodium dodecyl sulfate (sds), by Merck Group (1 mentions)
Pluronic f 127, by Merck Group (1 mentions)
L lysine, by Merck Group (1 mentions)
Tween 20, by Merck Group (1 mentions)
Clone r35 95, by BD (1 mentions)
H perfluorooctyl silane, by Merck Group (1 mentions)
Evans blue dye, by Merck Group (1 mentions)
Phosphate buffered saline (pbs), by Thermo Fisher Scientific (1 mentions)
Sylgard 184, by Dow (1 mentions)
Dichloromethane, by Merck Group (1 mentions)
Yellow dual cartridge, by RS Components (1 mentions)
Perfluorohexane, by Apollo Scientific (1 mentions)
Multisizer 3, by Beckman Coulter (1 mentions)
Lecithin, by Sinopharm (1 mentions)
10 protocols using polyoxyethylene 40 stearate
1
Preparation of Naproxen-Amino Acid Complexes
2
Biotin-Targeted Microbubble Synthesis
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Lipid-shelled decafluorobutane MBs with functionalized biotin linker moieties were prepared by sonication of a gas-saturated aqueous suspension of distearoylphosphatidylcholine (2 mg/mL, Avanti Polar Lipids), polyoxyethylene-40-stearate (1 mg/mL, Sigma-Aldrich), and 1,2-distearoyl-sn-glycero-phosphoethanolamine-N-[biotinyl(polyethylene glycol)-3400] (0.14 mg/mL, Creative PEGWorks) in a glass vial. The MBs were washed by flotation-centrifugation to remove lipids not incorporated into the MB shell. MBs targeted to VCAM-1 (MBVCAM-1) were prepared by conjugation of biotinylated rat anti-mouse VCAM-1 antibody (Clone 429/MVCAM.A, Cat #. 553331, BD Pharmingen) to the MB surface using biotin-streptavidin-biotin linking. Control MBs (MBCtr,) bearing a nonspecific biotinylated rat IgG2a, K isotype control antibody (Clone R35-95, Cat #. 553928, BD Pharmingen) were also prepared.
3
Fabrication of Polymeric Microfluidic Devices
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Poly(dimethylsiloxane) (Sylgard® 184) was purchased from Dow Corning Corporation (Michigan, USA), and poly(methyl methacrylate) from theplasticshop.co.UK (Coventry, UK). Epoxy adhesive (Yellow Dual Cartridge) was purchased from RS Components Ltd. (Corby, UK). Poly(lactic-co-glycolic acid), Pluronic® F127, dichloromethane, polyoxyethylene (40) stearate, trichloro-(1 H, 1 H, 2 H, 2 H-perfluorooctyl)-silane, and Evans blue dye were purchased from Sigma Aldrich (Gillingham, UK). Phosphate buffered saline was purchased from Life Technologies (Thermo Fisher Scientific Inc., Massachusetts, USA) and 1,2-distearoyl-sn-glycero-3-phosphocholine from Avanti Polar Lipids (Alabama, USA). Perfluorohexane was purchased from Apollo Scientific Ltd. (Stockport, UK). Nitrogen (N2) gas was provided by The BOC Group plc (Guildford, UK).
4
Phospholipid-Encapsulated Microbubble Preparation
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For therapy, phospholipid-encapsulated MB containing perfluorocarbon gas (Perfluorobutane, Fluoromed, Round Rock, TX) were prepared in house and measured by electrozone sensing using a 50 μm aperture (Multisizer 3, Beckham Coulter, Brea, CA) 21 (link). In brief, 1,2-distearoyl-sn-glycero-3-phosphocholine (Avanti Polar Lipids Inc., Alabaster, AL), polyoxyethylene(40)stearate (Sigma Aldrich, St Louis, MO) and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000] (Avanti Polar Lipids) were mixed in chloroform in 88:11:1 molar ratios, dried under a stream of argon gas and stored under vacuum overnight. The film was rehydrated in 4 mL saline in a glass vial filled with an overhead of PFC. The lipids and PFC were sonicated for 75 s (XL2020, Qsonica LLC, Newtown, CT) and the resulting MB were washed 3 times in saline, yielding therapy microbubbles with a diameter of 3.5 ± 1.1 μm in diameter (Multisizer 3, Beckman Coulter) and were stored in sealed glass vials filled with PFC until usage (within 2 weeks of fabrication). Definity microbubbles (Lantheus Medical Imaging, North Billerica, MA) were used for contrast perfusion imaging. These MBs have a mean size of 1.1-3.3 μm and concentration of 1.2 × 1010 MB/mL and were administered via jugular access at a flow rate of 2 mL/h.
5
Targeted Microbubble Preparation for Cellular Binding
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Perfluorocarbon-filled, microbubbles with a lipid shell were prepared by sonicating an aqueous suspension of distearoyl phosphatidylcholine (2mg/ml; Avanti Polar Lipids, Alabaster AL, USA) and polyoxyethylene-(40)-stearate (1mg/ml; Sigma) that was gas-saturated. For targeting of microbubbles to leukocytes (MBLc), distearoyl-phosphatidylserine (0.3mg/ml; Avanti Polar Lipids) was added to the suspension before sonication [8 (link)]. For targeting of P-Selectin (MBPSel) and CD4 (MBCD4), distearoyl-phosphatidylethanolamine-PEG(3400)-biotin (0.14mg/ml; Creative PEG Works) was added to the suspension. Anti-P-Selectin antibody (RB40.34) or anti-CD4 antibody (H129.19) were then conjugated to the microbubble shell using biotin-streptavidin linking as previously described [9 (link)]. Control microbubbles (MBIso) with a non-specific control antibody of the same isotype (R3-34) were also prepared. We have previously shown that these protocols result in microbubbles of similar mean sizes of 2.5 to 2.7μm [7 ].
6
Lipid Microbubble Preparation Protocol
7
Preparation of Lipid Microbubbles
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Lipid MBs were prepared from a lipid aqueous dispersion composed of polyoxyethylene(40) stearate (Sigma-Aldrich; St. Louis, MO), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[bio-tinyl(polyethylene glycol)-2000] (DSPE-PEG2000-biotin) (Avanti polar lipids; Alabaster, AL), as previous described 30 . Briefly, polyoxyethylene (40) stearate, DSPC and DSPE-PEG2000-biotin (1/2/1, w/w/w) was dissolved in chloroform. The chloroform was evaporated by flushing with argon, followed by overnight vacuum-drying. The dried lipid film was rehydrated in 0.9% sodium chloride saline (final lipid concentration as 10 mg/mL) for 4 hr at room temperature. After a brief tip sonication to dissolve any lipid debris, the lipid dispersion was sonicated with a 20 kHz probe (Heat Systems Ultrasonics, Newtown, CT) in the presence of perfluorobutane gas (FluoroMed, L.P., Round Rock, TX). After sonication, the MBs were washed with 20 mL saline twice to remove any free lipid and were suspended in saline saturated with perfluorobutane. The lipid MBs were aliquoted in vials with perfluorobutane filled in head space and were stored at 4 ºC until use.
8
Preparation of Lipid-Shelled Microbubbles
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Following Mott et al. [26 (link)], lipid-shelled MB-PS were prepared by performing a sonication of a decafluorobutane gas-saturated aqueous suspension of 2 mg/mL distearoylphosphatidylcholine, 0.3 mg/mL distearoyl phosphatidylserine (Avanti Polar Lipids, Alabaster, AL, USA), and 1 mg/mL polyoxyethylene-40-stearate (Sigma-Aldrich, St. Louis, MO, USA). The fluorescent lipid-shelled MB-PS were prepared by adding 0.2 mg/mL tetramethylindocarbocyanine perchlorate (Sigma-Aldrich, St. Louis, MO, USA) or 0.2 mg/mL dioctadecyloxacarbocyanine perchlorate (Invitrogen, Waltham, MA, USA) to the gas-saturated aqueous suspension described above.
9
Cytotoxicity Assay for Cell Viability
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Etoposide (≥98%), dimethyl sulfoxide, MTT formazan, and polyoxyethylene (40) stearate were purchased from Sigma-Aldrich Co. (St Louis, MO, USA). Stearic acid, lecithin, Tween-20, and chloroform were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, People’s Republic of China). Deionized purified water was prepared in the laboratory and other reagents were all analytical grade. RPMI-1640, fetal calf serum, penicillin G, streptomycin, and trypsinase were obtained from Thermo Fisher Scientific (Waltham, MA, USA).
10
Microbubble Synthesis for Ultrasound Imaging
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