Beckman avanti centrifuge j 26xp

The HUCMSCs were subsequently seeded into a 6‐well plate at a density of 1 × 10⁵ cells/well and cultured in DMEM containing 10% FBS and 1% penicillin/streptomycin (Gibco, Grand Island, NY) until cell confluence reached 80%. The cells were then cultured for 48 hours with Roswell Park Memorial Institute (RPMI) 1640 medium containing EV‐depleted FBS (centrifuged at 100 000 g for 18 hours). The cell supernatant was then centrifuged using Beckman Avanti Centrifuge J‐26XP (Beckman Coulter) for debris and apoptotic body removal. The supernatant was subsequently centrifuged at 110 000 g for 70 minutes (Beckman Optima L‐80 XP Ultracentrifuge with 70Ti rotor), followed by purification by centrifugation at 110 000 g for 70 minutes. All the centrifugations were conducted at 4°C. The precipitates were then resuspended in PBS and sterilized by filtration through a 0.22‐μm filter (Millipore, Darmstadt, Germany).

hBMSCs were seeded into 6-well plates at a density of 1 × 10⁵ cells/well, and cultured in a DMEM medium containing 10% FBS solution and 1% Penicillin-Streptomycin (all from Gibco; Thermo Fisher Scientific, Inc., Waltham, MA, USA) until the cells reached 80-90% confluence. hBMSCs were cultured in a RPMI1640 medium containing EV‐depleted FBS (by centrifugation 18 h at 100,000 g) for 48 h. The supernatants were centrifuged at 2000g for 10 min and then 10,000g for 30 min to isolate the debris and apoptotic bodies (Beckman Avanti Centrifuge J‐26XP, Beckman Coulter, Brea, CA, USA) from the supernatants. Subsequently, the supernatants were centrifuged at 110,000g for an additional 70 min (Beckman Optima L‐80 XP Ultracentrifuge with 70Ti rotor), followed by washing with PBS solution and purification by centrifugating at 110,000g for 70 min. All the centrifugations were performed at 4 °C. The pellet was resuspended in PBS solution and sterilized by filtration through a 0.22 μm filter (Millipore, Darmstadt, Germany).