For human samples, venous blood was collected and the serum was separated by centrifugation at 700 × g and stored at −80°C until analyzed. The cytokines in human serum were analyzed by CBA. Briefly, IL-1β, IL-6, IL-8, MIP-1β, TNF-α, and MCP-1 concentrations of CB workers and control workers were measured. Human Soluble Protein Master Buffer Kit, Human IL-1β Flex Set (Bead B4), Human IL-6 Flex Set (Bead A7), Human IL-8 Flex Set (Bead A9), Human MCP-1 Flex Set (Bead D8), Human MIP-1β Flex Set (Bead E4), and Human TNF Flex Set (Bead C4) were used according to the manufacturer’s protocol (BD Biosciences). Data were acquired on BD Canto and BD LSRFortessa flow cytometers and analyzed by FCAP Array Software (Soft Flow Inc., Pecs, Hungary).
For animal samples, the blood was collected from angular vein after deep anesthetization with chloral hydrate. The angular vein is a small vein near the eye. The angular vein was pierced by a needle and the blood was collected. Blood and lung homogenate were centrifuged at 700 × g and then the supernatants were collected and stored at −80°C until analyzed. IL-6, IL-8, IL-1β, MIP-1β, and TNF-α were evaluated with commercially available mouse ELISA kit according to manufacturers’ recommendations (IL-6, IL-1β, MIP-1β, and TNF-α kits were from R&D Systems, Minneapolis MN and IL-8 kit was from Cusabio Biotech Company, Wuhan, China).
For animal samples, the blood was collected from angular vein after deep anesthetization with chloral hydrate. The angular vein is a small vein near the eye. The angular vein was pierced by a needle and the blood was collected. Blood and lung homogenate were centrifuged at 700 × g and then the supernatants were collected and stored at −80°C until analyzed. IL-6, IL-8, IL-1β, MIP-1β, and TNF-α were evaluated with commercially available mouse ELISA kit according to manufacturers’ recommendations (IL-6, IL-1β, MIP-1β, and TNF-α kits were from R&D Systems, Minneapolis MN and IL-8 kit was from Cusabio Biotech Company, Wuhan, China).