Fs30d bath sonicator

To fabricate the nanoparticles, a previously reported membrane coating approach was employed.^{37 (link)} First, poly(lactic-co-glycolic acid) (PLGA; carboxy-terminated, 50:50, 0.67 dL/g; Lactel Absorbable Polymers) cores were prepared by precipitating the polymer dissolved at 10 mg/mL using acetone into water, followed by evaporation to remove the organic solvent. Human RBC (hRBC) membrane ghosts were obtained by the hypotonic lysis of human O-negative RBCs (BioreclamationIVT). Human RBC membrane-derived vesicles were prepared by brief sonication of hRBC ghosts using a Fisher Scientific FS30D bath sonicator. To make hRBC-NPs, the membrane and PLGA cores were mixed together, followed by sonication to induce membrane fusion. Size and zeta potential were measured by dynamic light scattering (DLS) using a Malvern ZEN 3600 Zetasizer. To visualize the nanoparticles, the hRBC-NPs were deposited onto a 400-mesh carbon-coated copper grid (Electron Microscopy Sciences), stained with 1 wt% uranyl acetate (Electron Microscopy Sciences), and imaged using a Zeiss Libra 120 PLUS EF-TEM transmission electron microscope.

The cell membrane from B16-WT, B16-OVA, and B16-CD80/OVA cells was collected according to a previously published protocol.^{[46 ]} Briefly, cells were suspended in a lysis buffer containing 30 mM Tris-HCl pH 7.0 (Quality Biological) with 0.0759 M sucrose (Sigma-Aldrich), 0.225 M D-mannitol (Sigma-Aldrich), and a cocktail of phosphatase and protease inhibitors (Sigma-Aldrich), followed by physical disruption using a Kinematica Polytron PT 10/35 probe homogenizer at 70% power for 15 passes. The membrane was separated by first centrifuging at 10,000 g and then centrifuging the resulting supernatant at 150,000 g to obtain a membrane pellet using a Beckman Coulter Optima XPN-80 ultracentrifuge. To prepare polymeric cores, 1 mL of poly(DL-lactic-co-glycolic acid) (50:50 PLGA, 0.67 dL/g, Lactel Absorbable Polymers) in acetone at 10 mg/mL was added dropwise into 1 mL of water and the mixture was placed under a vacuum aspirator to evaporate the organic solvent. Membrane coating was carried out by mixing the preformed PLGA cores with the membrane at the appropriate ratios and sonicating the mixture for 2 min in a Fisher Scientific FS30D bath sonicator.

PNPs were prepared using a previously reported sonication method.[23 (link)] Polymeric nanoparticle cores were prepared using carboxyl acid-terminated 0.67 dL/g 50:50 poly(_DL-lactic-co-glycolic acid) (PLGA; LACTEL Absorbable Polymers) in a nanoprecipitation process. A volume of 1 mL of a 10 mg/mL PLGA solution in acetone was added rapidly to 4 mL of water. The acetone was then allowed to evaporate under vacuum for 3 hours. PNPs were prepared by fusing platelet membrane onto PLGA cores via sonication using a Fisher Scientific FS30D bath sonicator at a frequency of 42 kHz and a power of 100W for 2 minutes. The size and zeta-potential of PNPs were measured by dynamic light scattering (DLS) using a Malvern ZEN 3600 Zetasizer. To study the morphology of PNPs by transmission electronic microscopy (TEM), samples were deposited onto a 400-mesh carbon-coated copper grid (Electron Microscopy Sciences) and negatively stained with vanadium (Abcam).

Murine J774 cells (TIB-67, American Type Culture Collection) were maintained in Dulbecco’s Modified Eagle Medium (Mediatech) supplemented with 10% fetal bovine serum (HyClone) and 1% penicillin–streptomycin (Gibco). The cells were harvested by directly scraping them off of the bottom of T-175 tissue culture flasks, and the cell membrane was derived by a combination of mechanical disruption and differential centrifugation using a previously reported method.⁴⁵ Polymeric nanoparticle cores were synthesized by precipitating carboxyl-terminated poly(lactic-co-glycolic acid) (PLGA, 0.67 dL/g, 50:50 monomer ratio; LACTEL Absorbable Polymers) dissolved at 10 mg/mL in acetone into water, followed by evaporation under a vacuum to remove the organic solvent. To fabricate the final MΦ-NPs, the cell membrane was coated onto the preformed PLGA cores by mixing the two at a membrane protein to polymer weight ratio of 1:1, followed by sonication in a Fisher Scientific FS30D bath sonicator.