The neutrophils were prepared from human blood from healthy donors (ESF Paris, France) as described in [47 (link)]. Briefly, 500 mL of blood was sedimented in 2% dextran solution for 40 min and centrifuged 400 x g 8 min. Dulbecco PBS was added to the pellets, and then the neutrophils were separated from lymphocytes and the red cells by centrifugation for 30 min at 400 x g on Ficoll solution. The red cells were further eliminated after their lysis by centrifugation for 8 min, 400 g, 4°C. The pellet resuspended in PBS pH 7.4 containing 340 mM sucrose, 7 mM magnesium sulphate, 1 mM PMSF, 0.5 mM leupeptin was sonicated in the 30% pulse mode at power pulses (6) in an ice-cooled beaker 6 times during 10 s with resting time of 1 min between the sonications (sonicator XL, Misonix inc.). Neutrophil membranes and cytosol were separated by centrifugation for 1h30 at 200 000 g at 4°C. The membrane fractions were resuspended, aliquoted and stored at -80°C for further experiments.
Sonicator xl
Manufactured by Bioventus
Sourced in United States
The Sonicator XL is a laboratory equipment device that uses high-frequency sound waves to agitate samples for various applications. It is designed to provide efficient and consistent sample preparation for a range of research and testing purposes.
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Lab products found in correlation
5 protocols using sonicator xl
1
Neutrophil Membrane Preparation Protocol
2
Paclitaxel-Loaded Maleimide-Functionalized PLGA Nanoparticles
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PLGA–NPs surface functionalized with maleimide-terminated PEG and loaded with paclitaxel and coumarin-6 or rhodamine were synthesized as described in an earlier publication [9 ]. Briefly, PLGA (30 mg) and coumarin-6 (100 μg) or PLGA (25 mg) and PLGA-rhodamine (5 mg) along with paclitaxel (5 mg) were dissolved in 1 ml of chloroform. An oil-in-water emulsion was formed by emulsifying the polymer–drug solution in 8 ml of 2.5% w/v aqueous PVA solution by probe sonication (18–24 W; Sonicator XL, Misonix, NY, USA) for 5 min over an ice bath. The diblock copolymer PLA–PEG–maleimide was dissolved in chloroform (200 μl) and added dropwise to the above emulsion with stirring. The emulsion was stirred for 16–18 h under ambient conditions followed by 1 h under vacuum to remove the residual chloroform. NPs were washed twice by ultracentrifugation (35,000 rpm for 35 min at 4°C, Optima XPN-80, Beckman, CA, USA) and reconstitution in DI water. The final NP dispersion was then stored at 4°C until conjugation reaction on the same day.
3
Neutrophil Membrane Preparation for Experiments
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The neutrophils were prepared from human blood from healthy donors (ESF, Paris, France) as described in [60] (link). Briefly, 500 mL of blood was sedimented in 2% dextran solution for 40 min. PBS was added to the pellets, and then the neutrophils were separated from lymphocytes and the red cells by centrifugation for 30 min at 400g on Ficoll. The red cells were further eliminated after their lysis by centrifugation for 8 min, 400g, 4 °C. The pellet resuspended in PBS pH 7.4 containing 340 mM sucrose, 7 mM magnesium sulfate, 1 mM PMSF, 0.5 mM leupeptin was sonicated in the 30% pulse mode at power pulses (6) in an ice-cooled beaker 6 times during 10 s with interval of 1 min between the sonications (sonicator XL, Misonix Inc.). Neutrophil membranes and cytosol were separated by centrifugation for 1 h 30 min at 200,000g at 4 °C. The membrane fractions were resolubilized, aliquoted and stored at −80 °C for further experiments.
4
Preparation of LNT/BUD-Loaded Nanoparticles
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A simple solvent evaporation method with some modifications was used to prepare LNT/BUD-NPs. In brief, 50 mg LNT was dissolved in 20 ml deionized water. Meanwhile, 5 mg BUD was dissolved in 2 ml acetone to prepare BUD solution. In order to improve solubility, the above solution and organic phase were sonicated by a probe sonicator (Sonicator XL, Misonix, Melville, NY, United States) for 5 min. Then, the BUD solution was dropped into the LNT solution at a slow, uniform rate with stirring. To prepare the LNT/BUD-NPs, this mix solution was stirred at 300 rpm at 25°C for 12 h to eliminate all acetone. Finally, LNT/BUD-NPs were stored at 4°C for further experiments.
5
Lactoferrin-Based Nanoparticles for Drug Delivery
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LF@EMO-NPs were prepared using a slightly modified dialysis [15 (link)]. In brief, 100 mg LF was dissolved in 40 mL ultrapure water, along with the dissolution of 10 mg EMO in 2 mL DMSO. Then 2 ml of EMO solution was dripped into 40 ml of LF solution. To improve the solubility of EMO, the above solutions were sonicated for 5 min using a probe sonicator (Sonicator XL; Misonix, Melville, NY, USA), kept on a magnetic stirrer for 30 min to ensure uniform distribution and optimum size reduction, and dialyzed in ultrapure water with a dialysis bag (MWCO 1000, Millipore, USA) for 8 h. LF@EMO-NPs were passed through a syringe filter (0.22 μm) to remove impurities and free drug.
HA/LF@EMO-NPs were prepared by electrostatic adsorption [26 (link)]. Briefly, 4 mg of HA was dissolved in 2 mL of ultrapure water, followed by dropwise addition of HA solution to the LF@EMO-NPs solution after filtration. After stirring for 30 min and filtering through a syringe filter (0.22 μm), HA/LF@EMO-NPs were lyophilized until further use. All the above steps were performed under light-proof conditions.
HA/LF@EMO-NPs were prepared by electrostatic adsorption [26 (link)]. Briefly, 4 mg of HA was dissolved in 2 mL of ultrapure water, followed by dropwise addition of HA solution to the LF@EMO-NPs solution after filtration. After stirring for 30 min and filtering through a syringe filter (0.22 μm), HA/LF@EMO-NPs were lyophilized until further use. All the above steps were performed under light-proof conditions.
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