Bio plex 2 wash station

PBMCs were seeded at a concentration of 10⁵ cells per well in a v-bottomed 96 well plate, and then 200 μL of RF10 medium containing the relevant concentration of compound was added. After 18 h of incubation, the supernatant was removed from the cells and frozen at − 80 °C. Quantification of cytokines was performed using a custom Milliplex Human Cytokine Magnetic Kit (Millipore) according to the manufacturer's recommendations. Wash steps were performed using a Bio-Plex II Wash Station (Bio-Rad), and the data was acquired using a Bio-Plex 200 System with high-throughput fluidics (Bio-Rad). The resultant data was analyzed using Bio-Plex Manager 6.1 software.

Quantification of the levels of the murine cytokines IL-2, IL-6, MIP-1-alpha, and TNF-alpha in mouse plasma samples was performed using a custom Milliplex Mouse Cytokine/Chemokine Magnetic Kit (Millipore) according to the manufacturer’s recommendations. Wash steps were performed using a Bio-Plex II Wash Station (Bio-Rad), and the data were acquired using a Bio-Plex 200 System with high-throughput fluidics (Bio-Rad). The resultant data were analyzed using Bio-Plex Manager 6.1 software.